course 2 Flashcards
Oxidation
Decrease of electron density on carbon atom
Formation of chemical bond: C-O C-N C-X
Breaking chemical bond: C-H
oxidation means stealing electrons from carbon
Reduction
Increase of electron density on carbon atom
Formation of chemical bond C-H
Breaking chemical bond: C-O C-N C-X
Reduction means giving electrons to carbon
Phosphatase catalyzes
removing of Pi from a substrate
hydrolysis of an ester bond
Alanin aminotransferase (ALT)
catalyzes a reaction of Ala with
α-ketoglutarate. The reaction produces
Glutamate and pyruvate
Enzyme catalyzing cleavage of a bond
between 2 amino acids in proteins
belongs among
peptidases
hydrolases
What is Km?
KM = concentration of a substrate needed to
reach ½ Vmax of the reaction
KM = concentration of a substrate needed for transformation of ½ enzyme molecules to
complex enzyme-substrate
how can we tell if a reaction has a chance of occurring or not?
If a reaction can occur or not is determined by Gibbs energy
- Only reactions with negative Gibbs energy can take place
o At the same time, however, they need enough energy to surpass the
activation barrier – this is where enzymes help out
What is the kinetic equation?
v = k * [A] * [B] // k is coefficient, [A] is concentration A, [B] is concentration B
o Applies for reactions of I. and II. order
o Reaction of I. order are monomolecular reactions – one molecule spontaneously disintegrates into 2 molecules
▪ v = k * [A]
o reaction of II. order are bimolecular – two molecules react
▪ v = k * [A] * [B]
▪ a collision of a maximum 2 particles is assumed, a more complex reaction takes place via intermediates
Activation energy and temperature
v = k * [A] * [B]
o k = proportionality constant = k(-EA/RT) – the so-called Boltzmann factor
▪ expresses the fraction of molecules in
the system with energy higher than the
activation energy (EA)
- Boltzmann factor calculates which molecules have enough temperature to overcome the activation energy
- Effect of temperature – increasing the temperature
by 10 °C increases the reaction by 2-3 x
What are isoenzymes?
enzymes having the same function but different structure (and hence physical and chemical properties)
o if two different isoenzymes catalyse the same reaction, same product is formed
How is the rate of a catalyzed reaction determined?
using the michaelis Menten equation
What is Km?
is Michaelis constant - expresses the concentration of the substrate at which the reaction will proceed at half
maximum speed; that is, how much substrate is needed to feed 50% of the enzymes
o determines the affinity of the substrate to the enzyme - the higher the constant, the lower the affinity
o units - mol/l (because it expresses concentration)
Metabolism of macronutrients
macronutrients = carbohydrates, lipids, proteins
- carbohydrates and lipids break down to CO2 and H2O
- proteins break down into CO2, H2O and NH3
o NH3 is toxic to the body (mainly the brain) and is therefore converted to urea and excreted through urine
- all macronutrients have their own catabolic and anabolic pathways and some pathways in common (e.g Krebs cycle)
o their pathways are linked through carboxylic acids
catabolic reactions
glycolysis - degradation of carbohydrates to pyruvate / lactate
- glycogenolysis - degradation of glycogen to glucose
- lipolysis - degradation of triglycerides to glycerol + fatty acids
- beta-oxidation - degradation of fatty acids to acetyl-CoA
- ketone body breakdown – in case of starvation, liver produces ketone bodies which are send off to other tissues for
degradation
- degradation of proteins and amino acids
Anabolic reactions
gluconeogenesis – glucose synthesis
- glycogenesis – glycogen synthesis
- Fatty acid synthesis
- lipogenesis – synthesis of TAG from fatty acids and glycerol
- ketogenesis – production of ketone bodies in liver
- proteosynthesis – protein synthesis
- ornithine cycle – urea formation
Amphibolic reactions (some anabolic and some catabolic)
pyruvate dehydrogenase reaction – converts pyruvate to Acetyl-CoA
- Krebs or Citrate cycle - consumes Acetyl-CoA to produce CO2, NADH and FADH2
o note. NADH should be correctly written as NADH + H+
- electron transport chain - consumes NADH and FADH2 to produce ATP and H2O
carboxylic acids
pyruvate dehydrogenase reaction – converts pyruvate to Acetyl-CoA
- Krebs or Citrate cycle - consumes Acetyl-CoA to produce CO2, NADH and FADH2
o note. NADH should be correctly written as NADH + H+
- electron transport chain - consumes NADH and FADH2 to produce ATP and H2O
Hydroxy acids x keto acids
in the body, hydroxy acids are constantly converted to keto acids and keto acids to hydroxy acids
- a classic example is lactate (hydroxyacid) and pyruvate (keto acid)
conversion of hydroxy acid to keto acid is oxidation
–hydrogen is removed
o NAD+ is needed to take both hydrogens and becomes a NADH + H+
Conversion of keto acid to hydroxy acid is reduction
– hydrogen is added
o NADH + H+ is needed, which donates both of its hydrogens and becomes a NAD+
Detoxification of ammonia
transamination -> hydrolytic deamination / oxidative deamination -> urea synthesis
- everything takes place in the liver, transamination can take place elsewhere, urea synthesis also takes place in the
kidneys
- amino acids are degraded to CO2, H2O and NH3
what is the difference between an amino acid and a keto acid?
amino acids have NH3 and keto acids have oxygen
Transamination
amino acid and keto acid exchange NH3 and
oxygen
o AMK1 + KK2 -> KK1 + AMK2
o Coenzyme of transamination is pyridoxal phosphate
o alanine + α-ketoglutarate -> pyruvate + glutamate
o aspartate + α-ketoglutarate -> oxaloacetate + glutamate
Detoxification of ammonia
hydrolytic deamination - glutamine -> glutamate + NH3
- oxidative deamination - glutamate -> α-ketoglutarate + NH3
- urea synthesis (ornithine cycle) - CO2 + 2x NH3 -> urea
Decarboxylation
removal of the carboxyl group in the form of CO2
- amino acids become biogenic amines
o histidine -> histamine, hormone and neurotransmitter
o glutamate -> GABA (gamma-Aminobutyric Acid), inhibitory neurotransmitter
o tryptophan -> 5-hydroxytryptofan -> serotonin, one of the hormones of happiness
Carboxylation
opposite of decarboxylation - CO2 is added
- for example, in the synthesis of fatty acids
Dehydrogenation
oxidation of single bond to double bond
o -CH2-CH2- -> -CH=CH- + 2H+ + 2e-
o very frequent reaction in metabolism - for example in β-oxidation of fatty acids, Krebs cycle and synthesis of
unsaturated fatty acids
dehydrogenation of alcohols
primary alcohols
▪ alcohol -> aldehyde -> carboxylic acids
● methanol -> formaldehyde -> formic acid
● ethanol -> acetaldehyde -> acetic acid
▪ methanol itself is completely harmless, only after its metabolism to formic acid it becomes extremely toxic
all alcohols are metabolized by one enzyme (alcohol dehydrogenase),
secondary alcohols
▪ alcohol -> ketone
o tertiary alcohols – dehydrogenation does not occur
esterification
glucose could theoretically leave at the moment of entry into the cell, therefore in all cells immediately after entry it is esterified with phosphate to glucose-6-phosphate, which no longer crosses the membrane
o Only hepatocytes have an enzyme that can separate the phosphate again
oxidation
oxidation of glucose on 6th carbon -> glucuronic acid
o oxidation of glucose on 1st carbon -> gluconic acid
epimerization
fructose, mannose and galactose can be converted to one another though glucose
Is catalytically active RNA an enzyme?
catalytically active RNA is not an enzyme but a ribozyme
enzyme binding
– what substrate the enzyme works with
▪ absolute - the enzyme processes only one molecule (eg urease)
▪ relative - the enzyme processes one group of molecules (eg Hexokinase - phosphorylates all 6C sugars)
cofactors
protein part of a complex enzyme is apoenzyme, non-protein part is cofactor
o cofactors are
▪ organic – e.g. haeme
▪ inorganic - metal ion - Zn, Cu, Fe, Mn
● stabilize the active centre, aid redox reactions and polarize bonds
o division of cofactors
▪ prosthetic groups - firmly bound to the enzyme, part of the stable structures
▪ coenzymes - only weakly bound to the enzyme, can completely detach
Oxidoreductases - Aox + Bred -> Ared + Box
catalyze intermolecular oxidation-reduction reactions; transfer of hydrogen, electrons, or reaction with oxygen
- types of enzymes - dehydrogenases, oxidases, peroxidases, oxygenases
- eg alcohol dehydrogenase, lactate dehydrogenase (lactate <-> pyruvate), phenylalanine hydroxylase (phenylalanine ->
tyrosine)
what are coenzymes of oxidoreductases?
NAD (nicotinamidadenine dinucleotide) and NADP ((nicotinamidadenine dinucleotide phosphate)
FAD a FADH2 - flavinadenine dinucleotide
derived from riboflavin (vitamin B2)
coenzyme Q - ubiquinone/ubiquinol
▪ part of the respiratory chain
▪ prosthetic group – haem
Transferases - A-x + B -> A + B-x
transfer groups (-CH 3, -NH 2, phosphate) from the donor to the acceptor
- types of enzymes - C-transferases, glycosyltransferases, aminotransferases, phosphotransferases (or kinases)
what are the coenzymes of transferases?
ATP (adenosine triphosphate), GTP (guanosine triphosphate)
▪ carry phosphate groups
CoA - Coenzyme A
▪ transmits acyls
o TDP - thiamine diphosphate (also TPP - thiaminpyrophosphate)
▪ carries carbon groups
▪ precursor - thiamine (vitamin B1)
o PALP - pyridoxal phosphate
▪ carries the NH2 group
▪ precursor - pyridoxine (vitamin B6)
o THF - tetrahydrofolate
▪ carries single-carbon residues; precursor: folic acid
Hydrolases - A-B + H2O -> A-H + B-OH
catalyse hydrolytic cleavage of the substrate = cleaves bonds with the help of water
- we divide them into groups
o Proteases - cleaves peptide bonds in protein and peptide molecules
o glucosidases - cleaves glycosidic bonds
o lipases - cleaves ester bonds in lipids
o phosphatases - remove the phosphate group
o amylases - cleaves bonds between glucose molecules in polysaccharides
no coenzymes
Lyases - A-B -> A + B
enzymes catalyzing bond decomposition in a different way than hydrolysis or oxidation
- double bonds or cyclic compounds are often formed
- can cleave (or introduce) small molecules, e.g. H2O, CO2 or NH3
- coenzymes similar to transferases
Isomerases - A -> A‘
catalyse reactions within a molecule of one substrate, move atoms (groups) from one carbon to another
- e.g cis-trans-isomerase, ribose phosphate-isomerase, epimerase (changes the orientation of OH groups)
- most often they do not contain coenzymes
Ligases - A-b + C -> A-C + b
they catalyse the synthesis of simple molecules to complex molecules
- often energy-intensive bonds with simultaneous energy consumption (mostly ATP -> ADP + Pi)
- often contain coenzymes of transferases
o biotin (vitamin H) - carboxylation
Water soluble vitamins
Vitamin B1 - thiamine
Vitamin B2 - riboflavin
Vitamin B2 - riboflavin
Vitamin B5 – pantothenic acid
Vitamin B6 - pyridoxine
Vitamin B7 = Vitamin H = coenzyme R = Biotin
Vitamin B9 – Folic acid
Vitamin B12 - cobalamin
Vitamin C - Ascorbic acid
Vitamin B1 - thiamine
involved in carbohydrate metabolism
- active form - coenzyme TPP (thiaminpyrophosphate)
- metabolic function - transfer of hydroxy-alkyl residues = oxidative decarboxylation
o eg, oxidative decarboxylation of pyruvate and α-ketoglutarate
- symptoms of deficiency - fatigue, convulsions, digestive disorders, nerve disorders, beri-beri nerve disease
- sources - cereals, yeast, lentils, offal, yolk, pork
Vitamin B2 - riboflavin
components of flavoproteins, enzymes involved in oxidative reduction processes (respiratory chain)
- active form – coenzyme FAD
- symptoms of deficiency - inflammation of oral corners, lips, damage to mucous membranes and skin, growth arrest
- sources - meat, milk, eggs, liver, yeast, beer
Vitamin B3 - nicotinic acid
Also known as niacin form NIcotinic ACid vitamIN
- Active form – nicotinamide nucleotides NAD and NADP
- symptoms of deficiency - convulsions, nervous disorders, pellagra disease
- sources - meat, fish, yeast, multigrain cereal, lentils
Vitamin B5 – pantothenic acid
basis of coenzyme A, contribution to protein synthesis and oxidation reduction processes
- symptoms of deficiency - nervous disorders, convulsions
- sources - meat, cheese, eggs, liver, yeast, lentils
Vitamin B6 - pyridoxine
part of enzymes involved in amino acid metabolism (transaminases)
o pyridoxal phosphate -> transamination and decarboxylation of AA
- symptoms of deficiency - disorders of haemoglobin production, inflammation of skin and mucous membranes, epileptic
inflammation
- sources - liver, whole grain cereal products, egg yolk, yeast
Vitamin B7 = Vitamin H = coenzyme R = Biotin
significant coenzyme, promotes cell growth and division
- symptoms of deficiency - skin diseases, anorexia, fatigue
- sources - eggs, liver, vegetables, yeast, formed by intestinal bacteria
Vitamin B9 – Folic acid
affects the amino acid metabolism necessary for the formation of red blood cells
- active form - tetrahydrofolate(THF) - transfer of monocarbon residues
o carries methyl residues and alters deoxyuridine phosphate
- symptoms of deficiency: disorders of protein synthesis, anaemia
- sources - eggs, leafy vegetables, yeast
Vitamin B12 - cobalamin
does not occur in plants, it is formed only in animals
- ensures normal haematopoiesis
- consists of a tetrapyrrole skeleton (similar to haemoglobin) with a cobalt atom attached inside
- metabolic function - transport of methyl groups
- symptoms of deficiency - anaemia, degeneration of spinal nerves
- sources - liver, meat, intestinal bacteria
Vitamin C - Ascorbic acid
Allows for iron absorption, formation of collagen and erythrocytes, promotes blood clotting, production of antibodies, is
an antioxidant
- oxidoreductase cofactor (electron donor)
- symptoms of deficiency - gingivitis, bleeding, decreased resistance to infections, scurvy - the most severe stage of
avitaminosis
- sources - vegetables (Brussels sprouts, peppers), fruits (blackcurrants, strawberries), potatoes, internal organs of animals
Fat-soluble vitamins
Vitamin A - retinol
Vitamin D – calciferol
Vitamin E - tocopherol
Vitamin K - phylloquinone
Vitamin A - retinol
component of visual pigment, important for epithelial formation, antioxidant
- active forms
o retinal - vision, carbohydrate transport
o retinoic acid - signalling molecule, ensures development, differentiation, growth
- symptoms of deficiency - night blindness, drying of cornea and conjunctiva, rough skin, stopping growth
- sources - liver, egg yolk, butter, cheese, sea fish fat, provitamin (β-carotene) in plant foods (carrots)
Vitamin D – calciferol
group of vitamins, most important - D3 (cholecalciferol) and D2 (ergocalciferol)
- controls the metabolism of calcium and phosphorus, promotes their absorption from the small intestine and bone
deposition
- effective form - hormone calcitriol
- hypervitaminosis - increase of calcium absorption, its deposition in tissues and formation of kidney stones
- symptoms of deficiency - softening and deformation of bones - rickets (rachitis)
- sources - fat of sea fish, butter, liver, egg yolk, also due to UV radiation of the skin
Vitamin E - tocopherol
antioxidant, supports the activity of the gonads
- protection of the organism against cancer (together with A and C)
- symptoms of deficiency - muscle weakness, vascular system disorders
- sources - vegetable oils, cereal sprouts
Vitamin K - phylloquinone
supports the process of blood clotting, promoting the synthesis of prothrombin in the liver
- active form - phylohidroquinone
- symptoms of deficiency - impaired blood clotting
- sources - leafy vegetables, made up of intestinal bacteria
Michaelis-Menten equation
how fast the reaction will proceed is described by the Michaelis-Menten model - it suggests that the reaction rate
depends on the substrate concentration (higher concentration = higher speed)
Enzyme + Substrate <-> Enzyme-Substrate complex -> Enzyme + Product
Maximum, speed is achieved when the enzyme is saturated - at an infinite substrate concentration
o in practice this speed can be achieved, it is a real value
Michaelis constant (KM)
substrate concentration at which half of the maximum reaction rate is reached
o the concentration of the enzyme does not matter, only the substrate concentration
o we can imagine that the constant describes the affinity of the enzyme to the substrate - the lower the constant,
the higher the affinity
Competitive inhibition
competitive inhibitors “compete” with substrate molecules for the active
site of the enzyme
o Successful binding results in a dysfunctional enzyme-inhibitor complex
o these inhibitors often resemble the substrate - they must fit in the
same place as the substrate, so they must look similar
o E + I <-> EI
- it is a reversible action
- by increasing the substrate concentration, it is possible to decrease the efficiency of inhibition- by increasing the
substrate concentration we decrease the chance that the enzyme will pair with the inhibitor
o Vmax will remain the same – at infinite concentration of substrate competitive inhibitors will not be effective
- the action of a competitive inhibitor increases KM, because we need more substrate for the same reaction rate
Uncompetitive inhibition
the uncompetitive inhibitor is in all cases allosteric (= another binding site on the enzyme other than the active site)
o binding of the inhibitor causes a conformational change in the enzyme which is then unable to bind its substrate
- the inhibitor binds to the enzyme completely regardless of whether or not the substrate is bound to the enzyme= the
concentration of the reactants does not affect the potency of the inhibition
- vmax decreases - we have less functional enzyme molecules
o the inhibitor does not look at the substrate concentration at all - even if we increase the substrate concentration,
there will always be some of the enzyme molecules in the solution that will be inhibited
- KM stays the same- the inhibitor only takes out several molecules of enzyme, it does not change its affinity to the
substrate
Non-competitive inhibition
Inhibitors that bind only to the enzyme-substrate complex
o they are often allosteric
o E + S + I -> ES + I <-> ESI
- this inhibition is often observed with enzymes that bind multiple substrates - it
works by binding the first substrate to the enzyme, causing a change in the
conformation of the enzyme and revealing an additional binding site to which the inhibitor subsequently binds
- the inhibitor is very poor at low substrate concentration - it does not have enough ES complexes to bind to
o high substrate = lots of ES complexes= a lot of opportunities for the inhibitor to bind to an enzyme
- substrate concentration does not affect the binding of the inhibitor to the enzyme-substrate complex
- vmax and KM decrease (enzyme appears to have a higher affinity for substrate)
What does allosteric mean?
another binding site on the enzyme other than the active site
Acetylcholinesterase (AChE)
AChE is an enzyme responsible for the hydrolytic cleavage (degradation) of acetylcholine in the synaptic cleft between two neurons
is a hydrolase and contains serine in its active site
acetylcholine is a minor but very important neurotransmitter found
on neuromuscular plates, in the CNS and in the PNS
o It plays a role in maintaining consciousness, attention,
memory formation and muscle signaling
o it is an ester of acetic acid and choline
What is the effect of Acetylcholinesterase (AChE)?
AChE causes cleavage of the ester bond between acetyl and choline - acetic acid and choline is formed
- if the neuron is stimulated and the acetylcholine is released into the synaptic cleft, it must also be removed -
acetylcholine cannot act permanently, it would cause hyperstimulation
o for this reason, we directly breakdown acetylcholine via AChE
▪ the enzyme attacks the carbonyl carbon (C=O)
Inhibitory acetylcholinesterase
amplify signal on synapses that use acetylcholine as a neurotransmitter (cholinergic synapse)
irreversible inhibitors=
carbachol and sarin – nerve gas
reversible inhibitors=
edrophonium and donepezil
goblet cells
separate gland cells (technically it is a gland consisting of one cell)
▪ forms mucus mucin
▪ are in the GIT, the airway, the conjunctiva in the upper eyelid…
exocrine glands
secretion is produced on the body surface either directly or through the duct
o secretion exits the cell through its apical surface
o sweat glands, mammary glands, sebaceous glands, salivary glands…
endocrine glands
secretion remains in the body, most often goes from the cell to the surrounding connective tissue and then into the bloodstream
o secretion from the cell via basal surface
o pancreas, pituitary, adrenal gland, thyroid gland…
amphicrine glands
combination of exocrine and endocrine glands
merocrine secretion
secretory granules are excreted by exocytosis
o secretion synthesis is continuous but its secretion is not - secretion is stored inside the cell
o The merocrine cell has a lot of RER, GA and secretory granules
apocrine secretion („reverse phagocytosis “)
the cell secretes secretions by cleaving the portion of the apical cytoplasm in which the secretions are located
o typical for the mammary gland - fat droplets accumulate at the apical pole and the whole piece of the cell breaks
apart
o frequent in lipid-secreting cells
Holocrine secretion
the cell secretes secretion so in a way that it disintegrates completely and disappears by apoptosis
o typical for sebaceous glands
Eccrine secretion
secretion is secreted by individual molecules through the cell membrane (either by itself or via carriers) and is passively followed by water on the basis of osmosis
o Hydrophilic and ionic secretion needs transporters to get through the cell membrane
o the result of eccrine cell activity is mucus - solution of secrete, water and ions isotonic with cytoplasm
▪ from this solution, the ions are gradually pumped back into the cells
Cystic fibrosis (eccrine secretion)
caused by a malfunction of the membrane transporter for Cl-
▪ the consequence of a higher concentration of chloride ions leads to excessive reabsorption of sodium from mucus
▪ because water follows sodium, mucus dehydrates and increases its viscosity
▪ too thick mucus explains the symptoms of cystic fibrosis - thin tubes (pulmonary alveoli, vas deferens) clog and excessive mucus density interferes with the ability of antimicrobial peptides, leading to frequent
infections (mainly respiratory)
goblet cells
mucin mucus production; is oppressed by other cells, hence the shape of the cup
▪ have RER, large GA, apical cytoplasm filled with secretory granules
o in some organs, the whole surface epithelium is composed of mucin-producing cells (stomach, uterus)
what is the difference between intraepithelial glands and extraepithelial glands?
intraepithelial glands
o are built directly into the epithelium
o they do not have an outlet/ duct system- they open directly to the
surface of the epithelium
extraepithelial glands
o are placed under the epithelium of origin
o consist of a secretory section and a duct
Structure of the secretory portion
tubular - tube shape, most common, mucinous glands (see below)
o acinar -round shape, narrow lumen
▪ for these glands, serous secretion is typical - proteins, enzymes
o alveolar - shape of a bladder, have a wide, well recognizable lumen
- compound type of secretory portion
o tuboacinous- the secretory compartment has the shape of a tube with
a round end
o e.g. submandibular and sublingual
o tuboalveolar- there is a wide lumen at the end of the tube
▪ e.g. mammary glands in lactation, sweat glands
What are the types of ducts in compound glands?
intralobular- surrounded by epithelial structures, inside the lobule, leads directly from acinus to interlobular duct
● intercalated duct – forms part of the intralobular duct
● striated- connects the intercalated duct to the interlobular duct. Also, part of the interlobular ducts
▪ interlobular- outlet between lobule
▪ main - connects to the interlobular ducts, it is the last duct before the secretion exits the gland
serous secretion
is produced by alveolar and acinous cells
is basophilic - most often it is peptide hormones, proteins and enzymes
o Serous secreting cell (aka serous cell) has round nucleus, RER, GA and secretory granules
mucinous secretion
is produced by tubular cells
o goblet cell - Produces mucus rich in mucin
o mucinous cell has an active nucleus, GER, GA, and a secretory granule
o mucin secretory granules are well stainable by PAS dye, which is sensitive to polysaccharides
mixed secretion
seromucinous
o both types of secretory compartments occur side by side and also in direct combination - e.g. tuboalveolar
o Serous semilumen - a cap from serous cells at the end of the mucinous tubule
What are myoepithelial („basket“) cells?
something between smooth muscle and epithelium
- contractile epithelial cells used to expel secretions from
secretory compartments and into ducts
- slender star-shaped cells (basket cells) which are found
in glandular epithelium as a thin layer above the basement
membrane but generally beneath the luminal cells
- they contain cytokeratin’s as epithelial cells and actin and
myosin as smooth muscle cells
Endocrine glands
reverse polarization - the cell secretes secretion towards the basalis lamina
- construction of endocrine glands - always trabecula of cells, only in the thyroid gland
we find follicles (due to storage of iodine)
DNES - Diffusion Neuro Endocrine System
- endocrine cells scattered everywhere
along the GIT, secrete signalling molecules to regulate digestive system function - types of endocrine glands according to hormones produced
o polypeptides and proteins - adenohypophysis, parathyroids, islets of
Langerhans…
o catecholamines - adrenal medulla
o steroid hormones - adrenal cortex, testis, ovary
o thyroxine – thyroid gland
Thyroid Gland
Cuboidal epithelium arranged around the follicle
- inside the follicle is colloid - thyroglobulin - a storage form of
thyroid hormones
Liver
amphicrine character
o Exocrine function - forms bile
o endocrine function - involved in the metabolism and production of plasma proteins
- the basic morphological unit of the liver is the lobule of the central vein - the hexagonal prism consists of the plates of hepatocytes
o The central vein sits right in the middle of the whole prism
o fenestrated capillaries and bile ducts lead from the central vein towards the periphery of the lobules
- at the point of contact of the three adjacent lobules there is a
triad - 1 vessel, 1 vein and 1 bile duct
Outer membrane of mitochondria
contains many porins - channels that allow molecules to pass through the membrane
o as a result, the outer membrane non-selectively permits all molecules smaller than 10 kDa
- Bcl proteins - part of outer mitochondrial membrane - important proteins in the regulation of apoptosis
- Contains a few enzymes of fatty acid metabolism and phospholipids
- unlike the cell membrane, it contains almost no cholesterol
Intermembrane space of mitochondria
similar composition to cytosol (much less protein than matrix)
- there are proapoptotic proteins, cytochrome C and a lot of protons
Inner membrane of mitochondria
the original membrane of the former prokaryotic cell
- almost impermeable and very selective - everything except for small uncharged molecules needs a carrier
- it incorporates the respiratory chain complexes
o ATP synthase forms elemental bodies on the inner mitochondrial membrane
- folds (cristae) - sometimes they are modified and form tubules (in cells producing steroid hormones)
- is composed of 75% proteins and 25% phospholipids
o contains specific phospholipids cardiolipins that help with membrane elasticity
Mitochondrial matrix
has a gel like consistency because of its high protein concentration (500 g protein per 1L matrix)
- contains enzymes of Krebs cycle, β-oxidation of fatty acids and ornithine cycle, various nucleotide coenzymes, inorganic
ions (Ca), mtDNA, relevant tRNA and mRNA, mitochondrial ribosomes, chaperones and chaperonins
Import of proteins into mitochondria
most proteins are imported from the cytoplasm - they are synthesized on ribosomes in the cytosol
- chaperones ensure that proteins remain unpacked so that they can be transported through the mitochondrial
membrane
- on the inner and outer membrane are translocators for proteins - Tom and Sam complexes external, Tim complexes
internal
o Tom (transporter of outer membrane) complex gets protein into intermembrane space
o Sam complex is for proteins to remain integrated into the outer mitochondrial membrane
o Tim (transporter of inner membrane) 23 complex pushes proteins from the intermembrane space into the
matrix
o Tim 22 complex integrates proteins from the intermembrane space into the inner mitochondrial membrane
Mitochondrial DNA = mtDNA
small (16.5 kbp) circular double helix without introns
- encodes 2 rRNA, 22 tRNA and 13 proteins
o most of the genome has moved into the nucleus of the host cell, making the mitochondria nucleus dependent
and unable to live independently - 600-1000 mitochondrial proteins are encoded by nuclear DNA
- in the matrix of mitochondria there is also a proteosynthetic apparatus and 70S ribosomes that can produce molecules
encoded by mitochondrial DNA
o mitochondrial ribosomes resemble bacterial ribosomes, which means that some antibiotics targeting bacterial
ribosomes may also affect mitochondria
What is the function of mitochondria?
production of ATP by oxidative phosphorylation
- Krebs cycle, β-oxidation, ketogenesis, steroid synthesis, ornithine cycle, gluconeogenesis, iron and calcium metabolism
(regulation of intracellular calcium, triggering of apoptosis or necrosis, synaptic plasticity and thermogenesis
- basolateral labyrinth - transfer of water and ions using Na / K ATPase with high consumption of ATP
o Insufficient activity of mitochondria leads to transmission disorders and thus to cell oedema
Apoptosis
programmed cell death requiring gene expression, proteosynthesis, and ATP
- triggers in response to apoptotic signal, stress, cell damage…
- mitochondria contribute by releasing proapoptotic factors from the intermembrane space, interrupting the supply of energy to the rest of the cell, and overproducing ROS (reactive oxygen species)
Defects in membrane permeability
membrane damage is one of the main causes of cell death
- the initial phase of cell damage is disruption of the membranes - their ability to selectively permeability is impaired
o this harms ATP production, leading to ATP deficiency, non-functional membrane pumps and cell swelling
Mitochondrial theory of aging
mitochondria naturally form oxygen radicals that may damage mtDNA or host cell DNA
- accumulation of mutations in mtDNA with age gradually decreases the function of mitochondria and respiratory
complexes
- mitochondrial function impairment affects long-lived cells - neurons, cardiomyocytes, muscle cells
- consequence - heart failure, muscle weakness, diabetes mellitus, dementia, neurodegeneration…
Mutations in mitochondrial DNA
maternal inheritance - inherited along the maternal line because paternal mitochondria are not passed on to the offspring
- most variable - there are several tens of copies of mitochondrial DNA and at the same time we have tens of thousands
of copies of mitochondrial DNA in the cell
- DAD - diabetes and deafness
o Mutated leucine gene synthesizes abnormal proteins
o damage of hair cells and pancreatic β cells
LHON - Leber’s hereditary optical neuropathy
o begins in middle age with optic nerve dying and ends with complete blindness
o can be caused by multiple mutations in complex 1 - mitochondria do not work well and their production of ATP is
not enough for powerful and energy-intensive nerves
Mutations in nuclear DNA (mitochondria)
onset of the disease in prenatal development or just after delivery
- inherited by Mendelian inheritance, because the mutated genes are found in the nucleus and not in the mitochondria
- Leigh syndrome - progressive degenerative brain disease
Therapy of mitochondrial diseases
with the help of classic medicines very difficult, practically at all
- possibility of so-called mitochondrial transplantation - the child then has 3 parents, from one egg, from the second
sperm and from the third mitochondria
o Legal only in UK
- Unfortunately, the CRISPR method does not work on mitochondria because the DNA can no longer cross the membrane
Division and fusion of mitochondria
there is a balance in the cell between fission and fusion mitochondria
- fusion - content mixing, protein replenishment, mtDNA repair, distribution of metabolic intermediates
- division - increase of mitochondria
Mitochondrial inheritance
mitochondria reproduce exclusively asexually
- mtDNA is inherited maternally - all mitochondrial embryo genetic information comes from the egg and not from the
sperm
- mtDNA mutates faster than nuclear genome - more susceptible to mutation accidents
o heart and nerve tissue (brain, retina) - most often mitochondrial diseases
o the cell is able to recognize and destroy mitochondria with defective mtDNA or trigger apoptosis
- mitochondria in the cell can exchange mtDNA but do not combine it
summary of glucose metabolism
C6H12O6 + O2 -> H2O + CO2 + Energy
during this time, the oxygen is reduced from 0 to -2, and carbon is oxidized from 0 to +4
o glucose degradation can conveniently be divided into two phases
▪ the first phase is the transfer of electrons to the transmitters (NAD, FAD) and the formation of CO2 – ie. glycolysis, Krebs cycle
▪ the second phase is the transfer of electrons to the final acceptor (oxygen) and the utilization of the energy we get
● Only in the second phase we need oxygen
● The process of the second phase of glucose metabolism is the so-called Respiratory Chain
Respiratory chain
„mitochondrial electron transport chain “
- is located in the inner mitochondrial membrane
- consists of 4 large enzymes (complexes), coenzyme Q and cytochrome C
Complex IV (cytochrome C oxidase; ferrocytochrome)
the last respiratory chain complex
- takes electrons from cytochrome C and passes them to oxygen
o oxygen is reduced from 0 to -2 = 4 electrons are needed for each O2
- cytochrome C - a relatively small protein containing one molecule of heme
o heme contains an iron atom that can pass between Fe2+ and Fe3+ - this is how electron transfer functions
o the whole cytochrome C can therefore carry only one electron at a time
o is electrostatically attached to the inner membrane from outside (not part of the membrane)
- complex IV is inhibited it cyanide
Complex III (ubiquinol-ferricytochrome C oxidoreductase)
transfers electrons from coenzyme Q to cytochrome C
- contains FeS (iron-sulphur) centre - an evolutionary ancient electron carrier
- coenzyme Q (ubiquinone) - electron transporter inside the inner mitochondrial
membrane
o it’s not a protein, it’s just a small molecule (on picture)
o has two forms - ubiquinone (oxidized form) and ubiquinol (reduced form)
o there are many other types of coenzyme Q - mammals have coenzyme Q10
▪ 10 because it contains 10 isoprene units
o contains two oxygen in its structure, which can be reduced and then re-oxidized - it can carry two electrons at the same time
▪ these two electrons can be obtained from many different sources - complex I, complex II, ETF dehydrogenase…
Complex I (NADH oxidase; NADH-ubiquinone oxidoreductase)
a giant complex composed of 45 subunits - it is much larger than the membrane itself
- oxidizes NADH + H+ to NAD+
o 2 electrons are released which travel to the coenzyme Q through complex I
Complex II (succinate dehydrogenase)
embedded in the inner mitochondrial membrane, but not completely - partially communicating directly with the
mitochondrial matrix
- active also in Krebs cycle – oxidizes succinate to fumarate
o In this reaction, 2 electrons are released and go through FAD to coenzyme Q
o FAD (unlike NAD) is not a separate molecule, it is just a part of enzymes (like a human hand)
Proton pumping
Complexes I, III and IV are large proteins that cross the entire inner membrane and pump protons from the mitochondria
matrix into the intermembrane space during electron transfer, creating a strong proton gradient
Complementary proteins of the respiratory chain
serves mainly as an alternative pathway of electron transport to coenzyme Q
- ETFDH (ETF dehydrogenase, electron-transferring-flavoprotein-dehydrogenase)
o ETF dehydrogenase contains FAD in its structure, thanks to which it can take electrons from ETF (electron transfer
flavoprotein) and then pass them on to coenzyme Q
- GPDH (glycerol- phosphate dehydrogenase)
o NADH, that is formed outside the mitochondria cannot spontaneously cross the mitochondrial membrane → it is
transmitted by GPDH and subsequently reaches the respiratory chain where it donates its two electrons
ATP synthase
an enzyme not part of the respiratory chain but also in the inner mitochondrial membrane
- whole protein works like a turbine
o has a proton channel that allows protons to pass from the intermembrane space to the matrix according to the concentration gradient
o As it passes through the proton channel, it turns ATP synthase - like water flowing through a hydroelectric power
station
- rotation of ATP synthase connects ADP and Pi into ATP
- oligomycin - bacterial product, ATP synthase inhibitor
Where does Krebs cycle occur?
occurs in the matrix of mitochondria
What is the main function of the Krebs cycle?
‘crossroad of metabolic pathways’ - the interconnection of carbohydrate, lipid, and protein metabolism
- amphibolic pathway (ie anabolic and catabolic pathway)
o catabolic function - energy source, ATP synthesis
o Anabolic function - formation of precursors for the synthesis of glucose, lipids, amino acids, porphyrins
- burns nutrients to the water, CO2 and reduced coenzymes that continue into the respiratory chain
Krebs cycle
1) synthesis of citrate (starting substance, hence citrate cycle)) - oxaloacetate + acetyl-coA -> citrate + CoA
2) dehydrogenation and hydrogenation -> formation of isocitrate (so called citrate activation)
3) dehydrogenation and decarboxylation -> formation of 2-oxoglutarate (or α-ketoglutarate)
▪ here CO2 is split off and NADH if formed
4) dehydrogenation, decarboxylation and binding of CoA -> formation of succinyl-CoA
▪ CO2 is slip off again and NADH is formed
5) cleavage of CoA -> formation of succinate
▪ GTP is formed
6) dehydrogenation -> formation of fumarate
▪ FADH is formed
7) hydration -> formation of malate
8) dehydrogenation -> formation of oxaloacetate
▪ NADH is formed
- total yield from 1 acetyl-coA = 2 CO2, 1 GTP, 3 NADH and 1 FADH2
o the use of cofactors in the respiratory chain is about 10 ATP
Regulatory enzymes of Krebs cycle
citrate synthase, isocitrate dehydrogenase and α-ketoglutarate dehydrogenase
They are enzymes that are inhibited by high concentrations of ATP, preventing excessive consumption of acetyl-CoA
pyruvate dehydrogenase or pyruvate dehydrogenase complex
an enzymatic complex that catalyzes the oxidative decarboxylation of pyruvate, which produces acetyl-CoA, which subsequently enters the Krebs cycle
● a number of coenzymes participate in this reaction along with the complex
▪ pyruvate + NAD+ + CoA -> acetyl-CoA + NADH + H+ + CO2
▪ pyruvate dehydrogenase reaction is irreversible
citrate synthase
catalyzes the first reaction of the Krebs cycle – condensation of oxaloacetate and acetyl CoA
o ATP is not required for the condensation of oxaloacetate and acetyl-CoA under citrate synthase, on the contrary -
if ATP is present, the Krebs cycle is generally inhibited (with sufficient energy, the cycle is not necessary)
aconitase
converts citrate into cis-aconitate (dehydration) and then cis-aconitate to isocitrate (hydration)
Isocitrate dehydrogenase
dehydrogenates isocitrate to α-ketoglutarate (= 2-oxoglutarate)
▪ the rate of reaction depends on the concentration of ATP and NADH -> excess of ATP or NADH means that the reaction will proceed slowly (again the excess of ATP and NADH indicates enough energy and therefore no need to produce more)
▪ this reaction generates NADH + H
α-ketoglutarate dehydrogenase
α-ketoglutarate -> succinyl-CoA + NADH + H+
▪ catalyzes oxidative decarboxylation with simultaneous binding of α-keto carbon to CoA and formation of NADH + H +
▪ the same mechanism as the oxidative decarboxylation of pyruvate
▪ coenzyme thiamine pyrophosphate is required
succinyl -CoA-synthetase
cleaves attached CoA and at the same time generates GTP (GDP+Pi -> GTP)
▪ stupid nomenclature of the enzyme, because it does not synthesize succinyl-CoA, but breaks it down
succinate dehydrogenase
Flavin enzyme with firmly bound zFAD and non-heme iron
▪ succinate dehydrogenase produces fumarate
▪ the reaction is carried out with simultaneous transfer of hydrogen to FAD, which is reduced to FADH2
▪ succinate dehydrogenase is also complex II in the respiratory chain
fumarate hydratase
hydrates fumarate into malate
malate dehydrogenase
dehydrogenates malate into oxaloacetate
▪ oxaloacetate and fumarate link the citrate cycle to the urea cycle
▪ oxaloacetate can also be used for glucose synthesis and aspartate (aspartic acid) synthesis
Sources of acetyl-coA
Coenzyme A is only a carrier of acetyl, it does not degrade during metabolism and can occur freely
- source of acetyl-CoA may be, for example, glycolysis occurring in the cytoplasm
o 1 glucose -> 2 pyruvates that enter the mitochondria and pyruvate dehydrogenase create acetyl-coA from them
(see above)
- β-oxidation of fatty acids occurring in mitochondria
o carries much more energy than glycolysis
- amino acids - degradation of some AA produces pyruvate,
others produce directly Krebs cycle intermediates
o alanine -> pyruvate; aspartate -> oxaloacetate…
What are cataplerotic reactions?
these are reactions that deplete the Krebs cycle intermediates
- if ATP is not needed, the Krebs cycle intermediates will begin to
convert to other useful compounds
o citrate - exits from mitochondria into the cytoplasm
where it cleaves to acetyl-CoA + oxaloacetate
▪ acetyl-CoA is used for the synthesis of fatty acids and steroids
▪ oxaloacetate returns to mitochondria or is transaminated to aspartate
o α-ketoglutarate -> its transamination forms glutamate (also the formation of glutamine, histidine …etc.)
o succinyl-CoA -> serves for porphyrin synthesis (=cyclic organic compound consisting of 4 pyrrole nuclei)
o malate, oxaloacetate -> either converted to AA (aspartate), or gluconeogenesis produces glucose
What are anaplerotic reactions?
anaplerotic reactions are reactions that produce citrate cycle intermediates
o typical example is the synthesis of oxaloacetate from pyruvate - pyruvate + CO2 + ATP -> oxaloacetate + ADP + Pi
▪ catalyzed by the enzyme pyruvate carboxylase with the participation of biotin cofactor
o metabolism of amino acids
▪ aspartic acid (Asp), asparagine (Asn) -> oxaloacetate
▪ glutamic acid (Glu), glutamine (Gln), histidine (His), proline (Pro), arginine (Arg) -> α-ketoglutarate
▪ valine (Val), threonine (Thr), methionine (Met) -> succinyl-CoA
▪ phenylalanine (Phe), tyrosine (Tyr) -> fumarate + acetylCoA
▪ alanine (Ala), serine (Ser), cysteine (Cys), glycine (Gly) -> pyruvate
o Fatty acids with an odd number of carbons -> after β-oxidation remains propionyl-CoA, which is converted to succinyl-CoA
How is Krebs cycle regulated?
is closely related to the respiratory chain, the consumption of ATP
and the consumption of reduced cofactors
- activation - low ATP/ADP or NADH/NAD
- inhibition - high ATP/ADP or NADH/NAD
- Krebs cycle regulatory enzymes = citrate synthase, isocitrate dehydrogenase and α-ketoglutarate dehydrogenase
Transport of metabolites (Krebs cycle)
the outer membrane of mitochondria has a lot of pores, which makes it quite permeable
o inner membrane tightly regulates the transport of substances and is almost impermeable -> carriers are used for the transport of larger molecules
- matrix is charged negatively (OH groups), intermembrane space charged positively (proteins)
▪ the membrane potential ranges from 180 to 200 mV
o molecules that can cross the membrane
▪ free diffusion - O2, CO2 a H2O
▪ antiport - pyruvate and H2PO4 for OH-, ATP for ADP, Ca2+ for 2H+
Shuttles
specific mechanism by which substances that would not normally pass through the inner mitochondrial membrane are transferred
Malate-aspartate shuttle
ensures the transfer of NADH from the cytoplasm to the mitochondria
1) in the cytoplasm NADH transfers hydrogen to oxaloacetate, resulting in malate formation
2) malate enters the matrix by antiport with α-ketoglutarate
3) malate passes hydrogen to NAD, oxaloacetate is formed again
4) transamination → oxaloacetate becomes aspartate and glutamate becomes α-ketoglutarate
5) aspartate enters the cytoplasm with the glutamate antiport
6) second transamination → aspartate becomes oxaloacetate and α-
ketoglutarate becomes glutamate
- result = in the cytosol less NADH and in the matrix more NADH
Glycerol phosphate shuttle
the way by which cytosol rapidly regenerates NADH to NAD in glycolysis
1) NADH in cytoplasm converts dihydroxyacetone phosphate to glycerol-3-phosphate
2) glycerol-3-phosphate transfers hydrogen to mGPD (mitochondrial glycerol-3-phosphate dehydrogenase)
o mGPDH has FAD as a prosthetic group, which then reduces coenzyme Q in the respiratory chain
- result = oxidized NAD in the cytoplasm and reduced coenzyme Q in
the respiratory chain
What are the characteristics of epithelial tissue?
Predominantly cells over extracellular matrix
o high adhesivety of cells
o polarization of cells
o avascular tissue - tissue nourishes by diffusion from the connective tissue under the basement membrane
o Strong nerve innervation
o high regenerative ability due to stem cells in the basement membrane
o originates from all 3 germ layers - ectodermal epidermis, endodermal glands, mesodermal endothelium
Diffusion barrier
one of the most important functions of the epithelium is to create compartments in the body and prevent free diffusion
o examples - intestine, kidneys, exocrine glands, brain capillaries (haematoencephalic barrier), plexus choroideus (barrier blood-liquor), thymus, testes
Junctional complex
zonula occludens (tight junction) cells leave no space between themselves
o serves for compartmentalization of surface plasmalema in resorptive epithelial cells
o zonula adherens (adhesive connection) - strong and mechanically resistant band connection using cell actin fibres
o macula adherens (desmosome) - strong and mechanically resistant point connection using intermediate filament cells
Zonula occludens
proteins claudins and occludins via adapter proteins connected to actin filaments of neighboring cells
o prevents the paracellular passage of substances => all substances must travel transcellularly
o permeability varies among different epitheliums
o impermeable - collecting ducts of kidneys, urothelium, capillaries in the brain
o almost permeable - small intestine, proximal tubulus of the kidney
Basement membrane
mediates interaction between cell and matrix o anchors cells
o attached to connective tissue via anchor fibrils
o consists of proteins and proteoglycans
o lamina basalis ensures attachment of cells to the basement
▪ lamina rara (lucida) - transmembrane proteins syndecan or integrins connect cell and rara
▪ lamina densa - it consists of collagen IV, laminin, perlecan
o lamina fibroreticularis - contains microfibrils and anchor fibers (fibrillin and collagen VI and VII), which fixate the basal lamina to the fibres of collagen III (connective tissue) under the basement membrane
o special basement membranes are made of two basement membranes (kidneys, lungs)
Types of epithelial transmission
simple diffusion - O2, CO2, NO
o passive transport (facilitated diffusion) - carriers, ion channels, aquaporins (for water)
o active transport - pumps (ATPases), the most important is Na / K ATPase
o for larger molecules - endocytosis (pinocytosis, phagocytosis), transcytosis (transport of substances from the apical
membrane to the basal or vice versa) and exocytosis
o aquaporins - transmembrane proteins with a hydrophilic tunnel that facilitate diffusion of water
o example in collecting duct in kidney, large intestine
Diffusion barrier in transporting epithelium
impermeable junctions - ileum and colon, water and ions must pass transcellularly (or controlled)
o permeable junctions - duodenum and jejunum, there is an absorption of ions and nutrients
o water passes paracellularly according to the concentration gradient
Pinocytosis
runs through pinocytic vesicles -> the cytoplasmic membrane invaginates and then gets excised
o some proteins trigger invagination of the plasmalema and the protein dynamin closes the sac
o clathrin-mediated endocytosis
o Clathrin coat - adapter protein
o the ingested molecule (LDL, transferrin) binds to the receptor
o Clathrin-independent endocytosis
o caveola vesicles - numerous on endothelial cells
o protein caveolin envelops the membrane
o functions - endocytosis of viruses, transcytosis, start of signalling
cascades (eg insulin)
Transcytosis
transfer of macromolecules across barriers - endothelium
o transmission of antibodies across the placenta into the fetus - transplacental transmission, passively from the mother’s body
o transmission of IgA antibodies through mucous membranes
Endothelium
vascular lining - barrier between blood and tissues
o simple squamous epithelium
o it has different permeability - from very permeable in bone marrow to impermeable in brain
o endothelial function
o gas exchange (diffusion), transport of substances, synthesis of vasoactive substances (mainly NO), vascular growth control, regulation of immune responses (facilitate the transfer of white blood cells to the site of inflammation) and participation in haemostasis
Transfer of substances through the endothelium
transcellular - blood-brain barrier
o paracellular - intestinal epithelium
o types of capillaries
o continuous - almost or completely impermeable
o fenestrated - the endothelium
has proteins that create holes and allow selective passage of substances
o sinusoid there are large holes in both the endothelium and its basement membrane that allow free passage of virtually all substances in the blood
Glykokalyx
glycoprotein and glycolipid layer, which covers the plasma membrane of some epithelium and other cells
o prevents blood clotting and white blood cells sticking
o allows cells of the immune system to pass to the sites of
inflammation
Blood - primary urine barrier
on one side is a fenestrated endothelium
o on the other hand, there are cytoplasmic processes of podocytes
o basement membranes of both are pivoted and pressed together -
one thick basement membrane is formed, which forms most of the barrier
o there is a lot of heparan sulphate in this membrane, which is negatively charged, which repels proteins (as they are also negatively charged) and helps keep them in the blood
o the gaps between the protuberances of the podocytes are quite small and the vast majority of proteins have no chance of getting through
o albumin is just below the permeability limit - if the kidneys do not work as they should, albumin will get through and can be detected in the urine - albumin thus acts as an excellent marker of kidney function
Blood-air barrier
thin-walled barrier - two epithelial cells (alveoli epithelium + endothelium) attached to each other, connected by basement membranes
o alveoli epithelium - single-layered flat epithelium (type I pneumocytes)
o continuous capillaries
The blood-brain barrier
BBB (Blood-Brain Barrier) - a barrier between the blood and the internal brain environment o several layers
o endothelium- continuous capillary
o basement membrane
▪ there may be pericytes in the basement membrane - a special cell type only for this occasion, helping to maintain BBB functionality and brain homeostasis
o astrocyte processes- supporting glial cells, play a role in BBB and neuronal sheath formation
o only water, a few gases and fat-soluble molecules can pass through the BBB, everything else needs carriers
o is due to well-formed zonulae occludentes and a small number of caveol
Multilayered epithelia as a barrier
multiple layers - basal, intermediate and superficial
o gradual cell differentiation - in the basal layer there are stem cells that proliferate and differentiate towards the surface
o tight junctions are present between the cells of the upper third of the epithelium (when they are up, they are not
needed below)
o thick basement membrane
o the basal layer is of cubic or cylindrical cells
o there is a fibrous layer under the basement membrane - either lamina propria or (in the skin) dermis
Layered squamous epithelium keratinised
- epidermis skin
o stratum corneum - lucidum - granulosum - spinosum - basale
o keratin is insoluble in water -> prevents water loss -> only lipophilic
substances pass through the skin
▪ in the stratum granulosum highest keratinization takes place
and there is also a large number of zonulae occludentes
▪ cytokeratin filaments and profilaggrin are involved in the production of keratin
Layered squamous epithelium non-keratinized
oral cavity, oesophagus, vagina, cornea, conjunctiva, larynx
o stratum basale - parabasale - intermedium - superficiale
o high glycogen content in superficial cells bacteria metabolize glucose anaerobically to lactate and ensure low pH
Melanocytes
protects against damage from UV radiation by melanin synthesis
o The melanocytes are present in the epidermis of the pars basalis
o melanin-containing granules are passed to cells in the stratum spinosum in which the granules disintegrate, and the
melanin is released into the cytoplasm
o melanosomes (organelles) - from GER and GA - start of melanin formation
o synthesis - dopa (enzyme tyrosinase) is synthesized from tyrosine, which polymerizes to melanin
Transient epithelium - urothelium
location - urinary tract
o lots of tight junctions
o surface cells (umbrelocytes) are the largest cells in the urothelium, they have differently bent membrane
o the rest of the epithelium - low cubic cells
o Special protein in tight junctions - uroplakin - even increases their effectiveness
o The surface of the cells (umbrelocytes) becomes more stained due to the presence of cytokeratin filaments
o uroplakin - in the membrane
o takes part in sealing as well
Use of monosaccharides in the body
glucose - energy production (glycolysis)
o energy supply - glycogenogenesis / fatty acid synthesis
o conversion to other monosaccharides - pentose cycle (to ribose)
o conversion to glucuronic acid by oxidation
o fructose - conversion to glucose
o galactose - conversion to glucose / lactose
o synthesis of glycoproteins and proteoglycans
Glucuronic acid
it is glucose with oxidized 6. carbon
- was first isolated in urine, hence its name from Urine
- biosynthesis proceeds by activation of glucose to UDP-glucose, followed by oxidation by enzyme UDP-glucose
dehydrogenase to UDP-glucuronic acid
- attaches to foreign hydrophobic substances in the liver to increase their solubility and to get out of the body via urine
- it further forms glycosaminoglycans, proteoglycans and proteoglycan aggregates
glycosaminoglycans (GAG)
These are long chains formed by alternating two molecules - uronic acid and hexosamine
uronic acids are acids containing both aldehyde and carboxylic acid (glucuronic)
hexosamines are hexoses with one OH group replaced by an amino group
o hyaluronic acid, chondroin sulphate, keratan sulphate, heparan sulphate, dermatan sulphate
proteoglycans (PG)
proteins (core proteins) to which all GAGs are perpendicularly bound (protein 5%, sugars 95%)
o fill the extracellular space and determine some properties - resistance to
pressure, return of tissue to its original shape, joint lubrication, articular
cartilage hydration, water retention, bone component
proteoglycan aggregate (PGA)
hyaluronic acid to which proteoglycans are bound perpendicularly through
binding proteins
Pentose cycle
alternative oxidative cleavage of glucose in the cell by NADP
o no ATP, NADH or FADH2 is produced
o the product of the pentose cycle is CO2, ribose-5-phosphate and 2 NADPH - the pentose cycle is the main source of
NADPH
o the pentose cycle can be divided into 2 phases - oxidation (irreversible) and monosaccharide mutual conversion
(reversible)
o cycle is regulated via NADPH - high NADPH / NADP ratio inhibits pentose cycle (NADPH is enough = no need to produce
another)
o enzymes for the pentose cycle are also present in erythrocytes, because NAPDH is used in defence against oxygen
radicals, which are much in erythrocyte - hexokinase (Fru -> Fru-6-P) and fructokinase (Fru -> Fru-1-P)
Oxidation phase of pentose phosphate pathway
at this stage, Glc-6-P is converted to ribulose-5-phosphate (Ru-5-P) and CO2 to form 2 NADPH
o dehydrogenation - Glc-6-P -> 6-phosphogluconolactate, the enzyme glc-6-P dehydrogenase, results in NADPH
o hydrolysis 6-phosphogluconolactate + water -> 6-phosphogluconate, enzyme 6-phosphogluconolactonase
o decarboxylation - 6-fosfoglukonát 6-phosphogluconate -> Ru-5-P + CO2, enzyme 6-phosphogluconate dehydrogenase,
NADHP is formed
Fructose metabolism
source of fructose in food - sucrose (disaccharide, glucose-fructose) from fruit
o fructose is activated by fructokinase (on Fru-1-P) or hexokinase (on Fru-6-P)
o Defective fructokinase disease is essential fructosuria and is essentially asymptomatic
o there is an aldolase B in the liver that can cleave Fru-1-P into glyceraldehyde (which is immediately phosphorylated to
glyceraldehyde-3-P) and DHAP, thereby forming glycolysis intermediates while skipping the major limiting factor of
glycolysis (a reaction catalyzed by phosphofructokinase 1) - fructose is thus metabolised much faster in the liver than
glucose
o defective aldolase B disease is hereditary fructose intolerance and is very dangerous
o fructose does not increase insulin production, its entry into cells is independent of insulin (using GLUT 2 and 5
transporters)
Sorbitol (glucitol)
a sugar alcohol produced by reducing the carbonyl group of fructose or glucose
o artificial sweetener E420
o glucose is naturally metabolized by the enzyme aldose reductase to sorbitol (NADPH is needed)
o important mainly in the liver, retina, lens, peripheral nerves and kidneys - sorbitol helps to maintain water
o there is a problem in patients with hyperglycaemia - changing osmolarity causes cataracts, peripheral neuropathy, renal and retinal damage and vascular problems
o sorbitol is further oxidized to fructose by the enzyme sorbitol dehydrogenase (NAD needed)
o important in liver and sperm (sperms get energy from fructose)
Galactose metabolism
dietary source - lactose (disaccharide, galactose-glucose)
o non-insulin-dependent entry into cells
o it is mainly metabolised to glucose in the liver
o phosphorylation- galactose + ATP -> Gal-1-P + ADP, enzyme galactokinase
o transfer UDP - Gal-1-P + UDP-Glc -> UDP-Gal + Glc-1-P, enzym uridyltransferáza
o isomerization- UDP-Gal -> UDP-Glc, enzyme epimerase
o galactokinase or uridyltransferase defect = galactosemia
o use - glycoproteins, glycolipids, GAG, breast milk lactose
Glucose
Central position in carbohydrate metabolism – all carbohydrates can be converted to glucose and vice versa
- Energy can be obtained from it even in the absence of O2
- All of our cells are able to use it and some tissues are even strictly dependent on it
o erythrocytes – because they do not have mitochondria they therefore do not
use Krebs cycle or the respiratory chain
o cells of CNS - however, during long-term starvation they adapt and 50% of their
consumption is covered by ketone bodies
Glycaemia = blood glucose concentration
normal fasting value is 3,3-5,6 mmol/l
o while after a meal it can be up to 7,1 mmol/l even in a healthy person
- regulation of blood glucose
o insulin – lowers glycaemia- glucose from blood enters cells
▪ cells use up glucose through glycolysis, pentose formation or
glycogen storage
o glucagon, adrenalin, growth hormone, cortisol – increase glycaemia
– glucose goes from liver to blood
▪ in hepatocytes, glucose is formed de novo by gluconeogenesis
or by degradation of glycogen storage
- sources of glucose - exogenous (food), glycogen breakdown and
gluconeogenesis (production of glucose from other metabolites)
Glucose transport across membranes
two different mechanisms
o secondary active transport using SGLT-1,2 (sodium-glucose transporter)
▪ glucose enters the cell by symport with sodium
▪ secondary active because the cell must expend energy to get rid of the sodium that came with glucose
▪ glucose is absorbed in the intestines and the proximal kidney tubule in this way
o facilitated diffusion through GLUT 1-7 transporters
▪ used in the transfer of glucose between blood and cells
What are GLUT?
GLUT are channels in the cytoplasmic membrane that can open and allow
glucose to pass freely across the membrane
▪ GLUT 1 - erythrocytes, blood-brain barrier
▪ GLUT 2 - liver, kidney, pancreatic β-cells, enterocytes
▪ GLUT 3 - brain
▪ GLUT 4 - adipose tissue, skeletal muscle, heart
● Insulin in the blood increases the amount of GLUT 4 transporters
Glucose phosphorylating and dephosphorylating enzymes
as soon as glucose enters the cell, it is activated by enzymes and ATP through
the phosphorylation from Glc to Glc-6-P - irreversible reaction
- Activation of glucose is the first step in its metabolism, while ensuring that
glucose does not flush out of the cell (phosphate cannot cross the membrane)
- two isoenzymes phosphorylate glucose
o glucokinase - in hepatocytes and pancreatic β-cells
▪ active at higher glycaemia (KM = 10 mM)
▪ β- cells respond to higher blood glucose levels via insulin secretion
o hexokinase - everywhere except hepatocytes and pancreatic β-cells
▪ active at much lower concentrations than glucokinase (KM = 0.1 mM)
▪ is inhibited by its product (Glc-6-P)
Dephosphorisation of glucose
converts Glc-6-P back to glucose by cleavage of inorganic phosphate
- is present only in the liver, kidneys and enterocytes
- is in smooth ER - Glc-6-P gets here via translocase
o is in this cell compartment so that newly formed glucose does not immediately phosphorylate in the cytosol
Glycolysis
catabolic reaction - conversion of glucose to 2 molecules of pyruvate (or lactate if we do not have enough oxygen)
- occurs in the cytoplasm of all cells with inorganic phosphate
- has two functions
o energy production under anaerobic conditions - during glycolysis in addition to pyruvate, ATP is produced directly
o source of acetylCoA - it can be used for everything possible
The first phase of glycolysis
First phase - glucose will be converted to fructose-1,6-bisphosphate (Fru-1,6-PP) with the investment of two ATPs
o 1. step – glucose activation - glucokinase or hexokinase enzyme, 1 ATP is consumed
2. step - isomerization - enzyme isomerase converts Glc-6-P to Fru-6-P
3. step - phosphorylation - 6-phosphofructo-1-kinase enzyme phosphorylates Fru-6-P to Fru-1,6-PP using 1 ATP
Second phase of glycolysis
Fru-6-P breaks down into two identical three-carbon monosaccharides
4. step - cleavage - the enzyme aldolase A divides Fru-1,6-PP into glyceraldehyde-3-phosphate (Gra-3-P) and dihydroxyacetone phosphate (DHAP)
DHAP is produced much more than Gra-3-P
5. step - isomerization - Triose phosphate isomerase enzyme converts DHAP to Gra-3-P
Third phase of glycolysis
Gra-3-P is converted to pyruvate, from which the cell obtains 4 ATP and 1 NADH
▪ since we now have two Gra-3-P molecules, the following reactions will run twice each
6. step - phosphorylation - enzyme glyceraldehyde phosphate dehydrogenase phosphorylates Gra-3-P to
1,3-bisphosphoglycerate (1,3-PP-Gly), inorganic phosphate (Pi) enters the reaction and NAD is reduced to NADH
7. step - dephosphorylation - phosphoglycerate kinase enzyme cleaves one phosphate from 1,3-PP-Gly to
form 3-phosphoglycerate (3-P-Gly) and one ATP
8. step - isomerization - phosphoglyceromutase enzyme converts 3-P-Gly to 2-P-Gly
9. step - dehydration - enzyme enolase from 2-P-Gly cleaves water, phosphoenolpyruvate (PEP) is formed
10. step - dephosphorylation - the pyruvate kinase enzyme cleaves the remaining phosphate, producing pyruvate and one ATP
- total 1 NADH and 2 ATP are extracted from glycolysis - two ATPs were invested in the first stage and two ATPs were obtained from each glyceraldehyde-3-phosphate in the third stage = -2 + 2 * 2 = +2
Metabolic fate of pyruvate
branch point of glycolysis
- fate of pyruvate depends on oxidative and redox state of the cell (sufficient O2 and NAD)
o aerobic conditions - pyruvate enters the MIT matrix, where it is converted to acetylCoA
o anaerobic conditions- pyruvate is converted to lactate (via enzyme lactate dehydrogenase) and released into the blood
▪ this happens because under aerobic conditions NADH is reduced in mitochondria, becomes NAD and can
again help with glycolysis
▪ under anaerobic conditions, however, NADH accumulates until finally there is no NAD and glycolysis stops
▪ when pyruvate is converted to lactate, NADH is converted to NAD and can immediately return to glycolysis
● Pyruvate + NADH + H+ <-> Lactate + NAD+
▪ there are no mitochondria in erythrocytes and so anaerobic glycolysis takes place there even under aerobic conditions
Regulation of glycolysis
the regulatory points are 3 enzymes
o glucokinase/hexokinase – activates glucose to Glc-6-P through using ATP
o 6-fosfofrukto-1-kináza (PFK-1) - phosphorylatesFru-6-P to Fru-1,6-PP through using ATP
▪ the main regulatory enzyme
▪ allosteric enzyme - the activator is Fru-2,6-PP (this molecule is produced by insulin)
▪ is inhibited
● if the cell has enough energy = high concentration of ATP or citrate in the cytoplasm, acidic pH
● in the presence of counter-regulatory hormones
o pyruvate kinase - dephosphorylates PEP to form pyruvate and ATP
Glycogenosis/Gluconeogenesis
apart of the regulatory points, gluconeogenesis is the same as glycolysis, but reverse
o glycolysis regulatory points are irreversible - gluconeogenesis bypasses them (bypass 1, 2 and 3)
- gluconeogenesis precursors - pyruvate, lactate, glycerol, oxaloacetate, propionate and glucogenic AMK (Ala, Gln…)
- takes place in the kidneys, liver and a little in enterocytes
- begins in mitochondria and then moves to the cytoplasm
- energetically consuming
o 2 Pyruvates + 4 ATP + 2 GTP + 2 NADH + 2 H+ + 4 H2O → glucose + 4 ADP + 2 GDP + 6 Pi + 2 NAD
Bypass 1 of gluconeogenesis
bypasses pyruvate kinase catalysed reaction (10th step of glycolysis)
- transport of pyruvate to mitochondria -> carboxylation of pyruvate (enzyme pyruvate carboxylase, cofactor biotin,
consumption of one ATP) -> formation of oxaloacetate (OAA) -> transfer of OAA to cytosol -> conversion of OAA to PEP
by PEP carboxykinase (consumption of GTP)
Bypass 2 of gluconeogenesis
bypasses PFK-1 catalysed reaction (3rd step of glycolysis)
- the enzyme fructose-1,6-bisphosphatase is used
- simply phosphate cleavage, no ATP formed, only inorganic phosphates
Bypass 3 of gluconeogenesis
bypasses the reaction catalysed by hexokinase (1st step of glycolysis)
- enzyme Glc-6-phosphatase (not found in skeletal muscle)
- Pi just breaks off again
Substrates for gluconeogenesis
Cori cycle
o Method by which erythrocytes and muscles get rid of lactate - they send it to the liver to turn it back into glucose
- alanine cycle
o the way energy is supplied to the muscles while also removing toxic ammonia
o Ammonia is linked to pyruvate by transamination, resulting in alanine that travels to the liver, where ammonia is
detached and processed into urea
o pyruvate is converted to glucose by gluconeogenesis and sent back to the blood
- amino acids
o all AA except Leu and Lys are able to degrade directly to pyruvate or Krebs cycle intermediates
- glycerol
o glycerol phosphorylation to glycerol-3-P - enzyme glycerol kinase
o dehydrogenation of glycerol-3-P to DHAP - enzyme glycerol phosphate dehydrogenase
Regulation of gluconeogenesis
active in glucose deficiency (starvation) and in pathological conditions (sepsis, polytrauma, burns, tumors …)
- gluconeogenesis is activated by stress hormones (counter-hormone hormones), inhibited by insulin
- major regulatory enzymes
o pyruvate carboxylase - activated by AcetylCoA
o PEP carboxykinase, Fru-1,6- bisphosphatase and Glc-6- phosphatase- regulated in the same way as glycolysis,
except vice versa
Energy Importance of Triacylglycerols (TAG)
o Lipids with energetic importance are called triacylglycerol = TAG
- store much more chemical energy than sugars and proteins - fats are hydrophobic, i.e. they do not bind water and are
not diluted by it (1g TAG is 100% fat, 1g glycogen is glucose + water is bound to it)
o 1g TAG has 6 times more energy than 1g glycogen
o supplies of glycogen and glucose are sufficient for one day, TAG for weeks
- the total weight of TAG in a healthy person is around 15% of the body weight
o the main site of TAG accumulation is the cytoplasm of adipocytes (fat cells)
Lipids as a source of energy
hormone-sensitive lipase (HSL) releases fatty acids from stores
o HSL performs complete lipolysis - 1 TAG -> 1 glycerol + 3 FA
o is inhibited by insulin and activated by counter-regulatory hormones (glucagon, adrenaline, noradrenaline)
- lipid utilization takes place in three steps
o lipid mobilization - hydrolysis of TAG to FA and glycerol and their transport by blood
o activation of FA in cytosol and their transport to MIT matrix
o β-oxidation - degradation of FA to acetyl-CoA, which enters the Krebs cycle
at the same time, reduced cofactors are produced, which are used directly in the respiratory chain
Mobilization of lipids
TAG is broken down into glycerol and FA and released from the cell into the blood
o FA longer than 12C need to have a carrier in plasma (because they are hydrophobic) - they bind to albumin
o glycerol is used in the liver (glycolysis / gluconeogenesis /…)
- conversion of glycerol to glycolysis intermediates
o Glycerol is the only component of TAG that can be converted to glucose through gluconeogenesis
first phosphorylation on glycerol-3-P using ATP (enzyme glycerol kinase)
then oxidation of glycerol-3-P to dihydroxyacetone phosphate using NAD + (enzyme glycerol-3-P dehydrogenase)
- entrance of FA into cells
o Short chain FA (<12C, do not need albumin) - they penetrate the membrane by simple diffusion
o Longer-chain FA’s use different transport systems in the membrane - this is facilitated diffusion
FATP - fatty acid transport protein
FAT/CD36 - fatty acid translocase
FA activation
involvement of FA in metabolism
- in cytosol FA is activated by CoA to acylCoA (enzyme acyl-CoA synthetase)
o FA + ATP + CoA -> acylCoA + AMP + 2Pi
o the resulting bond is between the acid and the sulphur atom CoA = thioester bond
Oxidation of fatty acids
β-oxidation occurs in the matrix of mitochondria and is the most common fate of activated FA (the so-called majority
pathway)
- ER membranes contain enzymes that catalyse ω- (omega) and α- (alpha) oxidation
o These oxidation pathways are called minority pathways, as they normally only consume about 5% of FA, but in
pathological situations their importance massively increases
o The Greek letters designate the carbon atom on which the reaction takes place
o take place in HER hepatocytes and tubular kidney cells on FA with 10-12C - dicarboxylic acids are formed
- short and medium FAs (below 12C) enter the mitochondria by simple diffusion
- the long chain (12-18C) must be activated and transferred using the carnitine chain
- very long (20C or more) enter the peroxisome where they are shortened
Entrance of FA into mitochodnrial matrix
acyl-CoA passes through the outer membrane easily, inner
membrane causes a little bit more trouble – carnitine and its translocases come into play here
1) FA leaves CoA and attaches to carnitine
2) carnitinacyltransferase I (CAT I) – on cytosolic side ofmitochondrial membrane
o transfers acyl from CoA to carnitine, resulting in acylcarnitine
3) acylcarnitine translocase - in the inner mitochondrial membrane
o Acylcarnitine is exchanged, via antiport, for
carnitine
4) Carnitinacyltransferase II (CAT II) - in matrix
mitochondria
o transfers acyl from acylcarnitine back to CoA, produces acylCoA
- free carnitine leaves matrix, via antiport, for acylcarnitine
β-oxidation of fatty acids
a cyclical process, repeating itself
- at each turn, the carbon chain is shortened by two carbons, which they release as acetyl-CoA
- one cycle has four steps
o dehydrogenation- acyl-CoA dehydrogenase enzyme
o hydration- enoyl-CoA hydratase enzyme
o dehydrogenation L-3-hydroxyacyl-CoA dehydrogenase enzyme
o thiolysis- β-ketothiolase enzyme
resulting reduced cofactors (NADH, FADH2) are used directly in the respiratory chain
Regulation of β-oxidation
works at the level of CAT I when FA enters MIT - is inhibited by malonyl-CoA (intermediate of FA synthesis)
o so called cross regulation- an intermediate of one metabolic pathway inhibits another metabolic pathway
o the principle of this regulation - in the synthesis of FA arises as an intermediate malonyl-CoA, which inhibits ß-
oxidation = it prevents the opposite processes in one cell (degradation x synthesis)
Animals cannot convert FA to glucose
FA are a rich source of energy for gluconeogenesis, but we do not get glucose from them (except for FAs with odd
number of C)
- AcetylCoA cannot be converted to pyruvate or oxalacetate - pyruvate dehydrogenase reaction is irreversible and both
carbons from AcetylCoA are cleaved off as CO2 during the Krebs cycle
- lants have another 2 enzymes in glyoxylate cycle - they can convert AcetylCoA to oxaloacetate
FA with an odd number of C
their β-oxidation is the same as in FA with even number of C, only two acetylCoA dont remain at the end, but one acetylCoA
and one propionylCoA
o PropionylCoA is converted to succinylCoA, which is involved in the Krebs cycle
succinyl CoA can be converted to oxaloacetate from which gluconeogenesis can synthesize glucose
- FA with an odd number of C is very little in the body
Degradation of unsaturated fatty acids
most unsaturated FA have a cis double bond configuration
- enoyl-CoA-hydratase requires only trans isomers -> it is necessary to convert cis isomer to trans by isomerase
- For the oxidation of unsaturated FAs, two additional enzymes are needed - isomerase and reductase
Oxidation of long chained FAs
takes place on peroxisomes (contain catalase)
o here is only a shortening -> then moving to mitochondria
- the pathway is induced by a high fat diet and hypolipiemic drugs – clofibrate
- oxidation ends with octanoyl-CoA, which is removed from peroxisomes by binding to carnitine
- the first stage is catalyzed by flavoprotein dehydrogenase - electron transfer to O2 -> H2O2
o is not attached to the respiratory chain -> FADH2 from the first step of β-oxidation is not reoxidised in the
respiratory chain, but in a reaction with O2
FADH2 + O2 -> FAD + H2O2
o peroxisomal catalase decomposes H2O2
2 H2O2 -> 2 H2O + O2
- reactions do not lead to the formation of ATP
Ketone bodies
there are 3 ketone compounds - acetoacetate, ß-hydroxybutyrate and acetone
o the first are the most essential, acetone is a more of a waste product and is useless
- can be completely degraded to CO2 and water
- derived from AcetylCoA - ketones are essentially a blood-transportable version of acetyl
- formed only in liver mitochondria
Function of ketones
they replace glucose as the main energy substrate during starvation - glucose runs out, so the liver begins to produce
ketone bodies and distributes them across the body to other tissues
- ketones can nourish all organs except liver (no enzyme) and erythrocytes (no mitochondria)
- AcetylCoA entry into KC depends on the availability of oxaloacetate - during fasting oxaloacetate is consumed for
glucose synthesis
o carbohydrate deficiency -> oxaloacetate deficiency -> KC slows down -> high AcetylCoA -> ketogenesis
o carbohydrate deficiency -> low glycemia -> activated HSL -> TAG breakdown -> high FA -> β-oxidation -> high
AcetylCoA -> ketogenesis
Synthesis of ketone bodies
- condensation- enzyme ketothiolase
- condensation- enzyme 3-hydroxy-3metylglutaryl-CoA synthase (HMG-CoA synthase)
- breakdown - enzymeHMG-CoA lyase
o HMG-CoA -> acetoacetate + AcetylCoA
o produces acetoacetate, which is the first ketone
in massive ketone formation β-hydroxybutyrate is
quantitatively the most important ketone in the blood
o diabetic ketoacidosis - a condition typical of type 1 diabetes mellitus
Ketone body activation
the liver produces them but does not use them – does not have CoA-transferase
o It would be unnecessary to synthesize ketone bodies if they were to be degraded immediately
- Once formed in the MIT matrix, ketone bodies enter the plasma
- one of the Krebs cycle intermediates - succinyl-CoA - is used to activate ketone bodies
- acetoacetate is activated by transfer of CoA from succinylCoA (using CoA-transferase) - succinate and acetoacetyl-CoA
are formed
- Acetoacetyl-CoA is then cleaved by β-ketothiolase to 2 acetyl-CoA
- β-hydroxybutyrate will first give its two extra hydrogens to the nearest NAD, thereby forming NADH and acetoacetate
Role of acetoacetate
cardiac muscle and kidney cortex prefer acetoacetate over glucose
- eventually the brain adapts to acetoacetate during starvation (up to 50% of
energy can be taken from ketone bodies)
- it also has a regulatory role - high levels of acetoacetate in the blood means
the presence of a large amount of acetyl-CoA, which is a signal to reduce
lipolysis (this means that the body has enough energy at that time)
Overview of material stored in the body
There is storage for fat (TAG) and saccharides (glycogen) but none for proteins (muscle breakdown is the last resort)
- TAG - convenient storage (because unlike glycogen it does not bind water)
What is the metabolism of glycogen like?
carbohydrate storage in animals, in the cytosol (explains its violet colour), in almost all cells
- in the liver (80-100g), skeletal muscle (300g) and in small quantities in each cell
- women have more glycogen - vaginal mucosa
o Liver glycogen - used to maintain stable blood glucose
o Muscle glycogen – used during short-term strenuous muscle work
o vaginal mucosa - glycogen -> lactate -> H + production -> acidic environment -> defence against microorganisms
- PAS reactions in histological preparations brightly colours glycogen violet
What is the structure of glycogen?
branched homopolymer
- Most glucose bound by α-1,4-O-glycosidic bonds
- branching from the strain of glycogen is by α-1,6 bond
- two groups of enzymes - one cleaves α-1,4 bonds, the other α-1,6 bonds
- glycogen has 2 ends - elongation and shortening takes place at non-reducing ends containing terminal glucose (enzymes
work with this end) - branching glycogen will provide more non-reducing ends - reducing end has a semi-acetyl hydroxyl – it is bound by the protein glycogenin, which forms the anchor at the beginning of the glycogen strain
What is glycogenesis?
all glycogen metabolism occurs in the cytosol
- 1) phosphorylation- enzyme glucokinase in liver and hexokinase in muscle
o glucose + ATP -> Glc-6-P + ADP
- 2) isomerization- enzyme glucose phosphate isomerase
o Glc-6-P -> Glc-1-P (= Cori ester)
- 3 coupling of UDP - enzyme UDP-glucose pyrophosphorylase
o Glc-1-P + UTP -> UDP-Glc -> formation of a macroergic compound
- 4 connection of UDP-Glc to non-reducing end of glycogen- enzyme glycogen synthase
o UDP is released and α-1,4 bond is formed
- the chain of linked glucose grows until it reaches a certain length and branches
- branching enzyme - branching enzyme - amylo-(1,4-1,6)-translocase
o catalyzes glycogen branching - it forms α-1,6 bonds
o new branches extended again by glycogen synthase
How is glycogenesis regulated?
MAIN ENZYME: glycogen synthase
regulation of glycogen synthase - phosphorylation (phosphorylated is inactive, dephosphorylated is active)
o activated by insulin (trying to lower free glucose levels in blood), inhibited by glucagon (trying to get more free
glucose into bloodi) and adrenaline
- phosphorylation of glycogen synthase is done by proteinkinase and dephosphorylation by phosphoprotein
phosphatase
What is Glycogenolysis?
- degradation of the glycogen molecule
- again, in cytosol
- phosphorolytic cleavage - cleavage of α-1,4 bond and formation of Glc-1-P (= Cori ester)
o Free inorganic P is used - linked to C1 glucose
o enzyme glycogen phosphorylase- cleaves α-1,4 bond – attaches phosphate to it
o ends 4 glucose molecules before branching
- phosphorolytic cleavage - cleavage of α-1,4 bond and formation of Glc-1-P (= Cori ester)
- isomerization Glc-1-P to Glc-6-P – enzyme phosphoglucomutase (glucose phosphate isomerase)
- Cleavage of branch – removal of α-1,6 bond
How is glycogenolysis regulated?
glycogen phosphorylase is active when phosphorylated - it is phosphorylated by phosphorylase kinase
- insulin inhibits phosphorylase kinase (insulin does not want to have more free glucose in the blood wants to prevent
glycogen breakdown) and counter-regulatory hormones activate it
- again, signalling via PKA and CAMK
How is malonyl-coA formed?
initial substance for the synthesis of FA is acetyl-CoA
- carboxylation to malonyl-CoA
o acetyl-CoA + ATP + HCO3 -> malonyl-CoA + ADP + Pi + H+
- enzyme acetyl-CoA carboxylase, biotin cofactor (or vitamin H / vitamin B7)
o regulatory enzyme of the whole pathway - also affects β-oxidation of FA
- CO2 removed during condensation with increasing FA
What are the requirements for palmitate formation?
8 acetyl-CoA, 14 NADPH and 7 ATP
- acetyl-CoA is transported from the mitochondria matrix using citrate
- 8 NADPH is obtained by transporting citrate to the cytoplasm and
the remaining 6 in the pentose cycle
How is fatty acid synthesis regulated?
sufficient substrates (carbohydrates / AMK) and energy are needed
o the main regulatory enzyme acetyl-CoA- carboxylase (active in dephosphorylated form)
o activated by – insulin and citrate – insulin and citrate - insulin activates glycolysis -> Acetyl-CoA, which is formed from
pyruvate after glycolysis, is required for FA synthesis
o inhibited by - glucagon, adrenalin, palmitoyl-CoA (if there is enough palmitoyl-CoA, there is no need to synthesize others) and AMP (in the absence of energy)
Elongation and desaturation of FA
elongation, enzymes elongase, if we need over 16C long FA
- formation of unsaturated FA is desaturation, enzymes desaturase
- both occur on the SER membrane on the side facing the cytosol
How does synthesis of triacylglycerols proceed?
final form of FA storage
- starting with DHAP (dihydroxyacetone phosphate, intermediate of glycolysis)
o reduction- enzyme DHAP dehydrogenase
o addition of the first two FAs - enzyme acyltransferase
glycerol-3-P + acylCoA -> monoacylglycerol-3-P + CoA
monoacylglycerol-3-P + acylCoA -> phosphate acid = diacylglycerol phosphate
o hydrolytic separation of phosphate from phosphate acid - enzyme phosphatidic acid phosphatase
phosphatidic acid + H2O -> 1,2-diacylglycerol + Pi
o connection of the last FA – again the enzyme acyltransferase
1,2-diacylglycerol + acylCoA -> TAG + CoA
What is muscle tissue?
tissue characterized by excitability, contraction and relaxation - due to myofilaments (actin and myosin II)
- Mechanocytes - cells capable of movement, have a high amount of actin and myosin
- types of muscles - smooth, striated skeletal and striated heart
sarco Greek “sarx” = meat
sarcolema - cytoplasmic membrane of muscle cells
o sarcoplasma - cytoplasm of muscle cells
o sarcomer - interval between two Z lines in muscle
o sarcoma - a tumour derived from muscle cells
myo - from Greek “mys, myos” = muscle
myocyte - muscle cell
o myofilaments - actin and myosin
o myofibril - a thin fibre inside muscle cells; it is a sarcomere chain attached at both ends to the sarcolemma
o myotoma - a block of embryonic cells from which muscles develop
o myocardium - muscle of the heart
What is a sarcomere?
skeletal and cardiac muscle
- five strips - A, I, M, H a Z
o A band indicates the presence of myosin
o I band indicates the absence of myosin, the
presence of actin
o M band indicates the centre of myosin fibers
o H band indicates gaps between actin fibers
o Z band indicates anchoring of actin fibers
- H and I strips shorten during contraction
- the sarcomere is the section between two Z strips
What is the contraction like in skeletal and cardiac muscle?
skeletal and cardiac muscle has large reserves of Ca2+ and is, therefore, less dependent on plasma calcium
o Ca2+ is released from the sarcoplasmic reticulum (that is a form of modified SER) into the cytoplasm where it binds to troponin C and activates myosin phosphatase - triggers myosin dephosphorylation (one of the contraction steps)
What is the contraction like in smooth muscle?
Ca2+ binds to the calmodulin, which then activates myosin light chain phosphorylating kinase
o Phosphorylation of light chain is required in smooth muscle for its binding to actin
in the dephosphorylated state, myosin is rolled up
Smooth muscle
contraction is not controllable by will - it is ensured by autonomous nerve
- consists of thin, spindle-like cells
o nucleus is centrally placed, long and narrow
o are full of actin, myosin, and intermediate filaments (desmin and vimentin)
o dense bodies (desmin) similar to Z lines; they are thickened sites where the filaments connect to each other or to the plasma membrane
- smooth muscle cells have cellular connections via hemidesmosomes and nex
o they transfer substances by pinocytic invagination
- occurrence - walls of hollow organs (thickness is about 150-200μm), blood vessels (15-20μm), skin, prostate eye
- smooth muscle contraction is induced by the mediator being spilled into the environment, transmitted through the nexes, automatically or stretched
Cardiac muscle
the basic building unit is a cylindrical cell - cardiomyocyte (85-100μm)
o lengthened cells, nucleus in the middle of the cell, a lot of mitochondria, a large supply of glycogen and lipids
o contain a lot of myoglobin - similar to haemoglobin but have only one binding site; is only in muscle cells
o diads - at the Z line level; it is formed by T-tubule (invagination of sarcolemma) and cistern of sarcoplasmic ER
What is an intercalated disc?
cardiomyocyte junction which contains several intercellular junctions
o fascie adherens - anchoring actin filaments
o desmosomes - anchorage of intermediate filaments
o nexes - they ensure the transfer of information between cells and serve to coordinate contraction
What is the cardiac conduction system?
an independent system that maintains the regularity and autonomy of heartbeat
o conduction system: sinoatrial (SA) node, internodal atrial connections, atrioventricular (AV) node, His bundle,
Tawar arms, Purkinje fibres
What are Purkinje fibers?
Purkinje fibres - contain much less myofibrils and mitochondria, lack of intercalated discs, have only nexes and local adhesive bonds; they contain a lot of glycogen
- cardiomyocytes have a very low ability to regenerate - in the case of cell death, a ligament is formed which is not
capable of conducting electrical excitement and cannot even contract
o this ligament can rupture under high pressure and blood starts to escape from the heart to the pericardium
cavity, where it accumulates and prevents heartbeat, which very quickly leads to death
Skeletal muscle
the basic unit of skeletal muscle is the fiber-multinucleated syncytium – (fusion of a large number of cells in one) with a
diameter of 60-100μm
- the skeletal fiber nuclei are oval and are located just below the sarcolemma
- muscle development - mesenchymal cells develop into myoblasts (which migrate and divide), which gradually begin to
connect to myotubules and to the final muscle fibers
skeletal muscles are covered by fascia (a string) on the surface, which is a cover of dense collagen ligament
What are satellite cells?
Satellites cells (belong to stem cells) serve for muscle growth and repair
- they are able to divide and fuse to prolong or repair an already existing muscle fiber
o their abilities are not unlimited, in case of too much damage the muscle dies and is replaced by connective tissue
- the sarcolemma in the skeletal muscle clings in and forms tubulous invaginations - so-called T-tubules
What is the sarcoplasmic reticulum?
sarcoplasmic reticulum (SER) - highly developed, modified ER full calcium forming a network around each myofibril
o together with invaginations of sarcolemma form so-called triads (SER-T tubules) at the A and I strip level
What are skeletal fibers that differ in structure, function and metabolism?
White fibre - larger; more actin, myosin and glycogen; fewer mitochondria, lipids and myoglobin
predominantly anaerobic metabolism - rapid
contraction of muscles, but quickly tired
suitable for sprints
o red fibre smaller; multiple mitochondria, lipids and
myoglobin; less actin, myosin and glycogen
mainly aerobic metabolism - slow muscle
contraction, but lasts for a very long time
suitable for marathons
What is a tendon?
collagen connective tissue that clamps the muscle into the bone
connective tissue coverage
endomysium - envelops one muscle fiber
o perimysium - envelops several bundles of fibers
o epimysium - envelops the entire muscle
How are myofibrils attached to the cytoskeleton?
protein dystrophin - inside stabilizes plasma membrane and connects it with actin (in Z bands)
o There may be many mutations in the dystrophin gene - mutations that damage the function of dystrophin (or its
auxiliary proteins) give rise to various diseases collectively called muscular dystrophy
o Duchenne muscular dystrophy - X-linked genetic disease where part of the dystrophin gene is deleted
Gradually laterally striated muscle breaks up, which eventually leads to death around the age of 20 due to respiratory failure
What are cellular movements based on?
Cellular movements are usually based on the interaction of motor proteins (molecular motors) with cytoskeletal structures – proteins move along cytoskeletal structures while using up ATP
What are molecular motors?
Kinesins: interact with microtubules
Dyneins: interact with microtubules
Myosins: interact with microfilaments
Kinesins
structure - 2 globular ATP binding heads (which interact with
microtubules), stalk and tail (made of light chains which bind cargo)
o movement uses up ATP
- function - transport of material into + end of microtubules (from centre to periphery)
- occurrence – mitotic spindle
Dynein
structure - 2 globular ATP binding heads (which interact with microtubules) and stalk (which binds cargo)
o head formed from 7 domains
- movement of dynein is quite chaotic and can be described as the walk of a
drunk sailor – movement forward follows is accompanied with wobbling from
side to side
- function - transport of material to – end of microtubule (from periphery to
centre)
- occurrence – mitotic spindle, flagella, cilia
Myosins
structure - globular ATP binding head (which interacts with actin microfilaments) and tail (which may bind cargo or bind to other myosins and form myosin filaments)
- types of myosin
o myosin-I – found in nearly all types of cells, monomer (1 head + 1 tail)
transport of vesicles inside the cell
o myosin-II – in muscles, dimers (2 heads + 1 tail in helix)
movement of the whole cell – contractile ring, muscle movement
- function of myosin – movement to + end of actin microfilaments
o transport of vesicle, movement of plasmatic membranes and muscle movement
Mitotic spindle
3 types of microtubules
o kinetochore which reach from centrosomes to kinetochores ( kinetochores are sites on centrosomes where microtubules anchor)
o polar – bind with polar microtubules of
opposing centrosome and push centrosomes
apart
o astral – reach out into all sides from
centrosome, bind to surrounding structures
and anchor the centrosome
Flagella and cilia
flagella – allows for cellular movement
- cilia – allows for cell to manipulate/move its surrounding
- flagella and cilia appear the same on the inside - 9 doublets of microtubules with a central pair of microtubules
o anchored by - end in basal body
- both structures move via the help of dynein – microtubule pairs are connected by dynein, which is anchored to one
microtubule and “walks” towards the other
o when microtubules are not tightly connected – one microtubule moves
o when microtubules and tightly connected – both microtubules bend towards the same direction
Intracellular transport of organelles
transport of organelles (along microtubules) is mediated by kinesin and dynein
- transport of vesicles (along actin microfilaments) is mediated by myosin I
Contractile ring and contractile bundles (actin filaments)
D – contractile ring - during cytokinesis at the equatorial line
“strangle” the cell
- B - contractile bundles - „muscle “of cells, attached to the
plasmatic membrane, can change shape of cell
- Myosin II helps with contraction
Amoeboid movement of cell
Used for example by macrophages
- Lamellipodia, filopodia, pseudopodia – projections of plasmatic
membrane filled with actin filaments
- growth of actin filaments and myosin II Æ growth of actin filament Æ
elongation of membrane Æ attachment of growth/projection to the
surface through focal adhesions Æ butt contraction (using myosin II)
- for amoeboid movement, you always need a stable surface
Muscle cells
structure – mainly myofibrils (actin microfilaments + myosin II)
- muscle contraction - myosin II drags actin filaments towards itself
o resting state – myosin bound to actin
o binding of ATP onto myosin causes actin to move away
o breakdown of ATP to ADP and Pi releases energy and myosin moves gently backward
o in its new location, myosin weakly binds to actin (onto a different monomer than before)
o Pi is released from myosin and the bound between myosin –actin becomes stronger
o in the last step, ADP is released which causes the movement of
myosin forwards which cause the movement of the whole actin
fiber
Muscle cells
structure – mainly myofibrils (actin microfilaments + myosin II)
- muscle contraction - myosin II drags actin filaments towards itself
o resting state – myosin bound to actin
o binding of ATP onto myosin causes actin to move away
o breakdown of ATP to ADP and Pi releases energy and myosin moves gently backward
o in its new location, myosin weakly binds to actin (onto a different monomer than before)
o Pi is released from myosin and the bound between myosin –actin becomes stronger
o in the last step, ADP is released which causes the movement of
myosin forwards which cause the movement of the whole actin
fiber
Proteins
Lack long-term stability, they constantly degrade and rebuild
- Protein daily turnover varies by tissue type- skeletal muscle 10%, liver 40%, small intestinal mucosa 80%
- Recommended daily intake (for 70kg male) – 100g
o Oxidation of 100g of protein will cover 10-20% of the bodies energy needs
Metabolism of amino acids
Sources of AAs – diet, degradation of body of body proteins, synthesis de novo
- Protein turnover is strictly regulated
- AA pool – amino acids, which are freely available in the body to be used, about 100g
o Highest level of AA in the body is glutamine and alanine – transport ammonia in blood
- Use of AA – proteosynthesis, degradation (to energy/ glucose/ FA), production of nitrogenous substances
o Excess AA cannot be stored (there is no storage protein) so it is burned as energy
