Energy Production: Carbphydrates Flashcards
What are the 4 stages in catabolism?
- Breakdown of fuel molecules to building block molecules
• Short pathways
• Breakage of C - N and C - O bonds (no C - C)
• No energy released - Degradation of building block materials to a small number of organic precursors
• Many pathways
• Small fraction of energy released • C - C bonds broken - Krebs cycle
• Carbon oxidised to CO2
• Small fraction of energy released - Electron Transport and Oxidative Phosphorylation
• Energy released
• ATP synthesised
What is stage 1 catabolism?
• Purpose - to convert nutrients to a form that can
be taken up into cells.
• Extracellular (GI tract)
• Complex molecules -> building block molecules
• Building block molecules absorbed from GI tract into circulation
• No energy produced.
What is catabolism stage 2?
• Intracellular
(cytosolic & mitochondrial)
• Many pathways (not all in all tissues)
• Building block molecules (many) -> simpler molecules (fewer)
• Oxidative (require coenzymes which are then reduced, e.g. NAD+ NADH)
• Some energy (as ATP) produced
What is catabolism stage 3?
- Mitochondrial
- A single pathway – Tricarboxylic acid (TCA) cycle
- Oxidative (requires NAD+, FAD)
- Some energy (as ATP) produced directly
- Acetyl (CH3CO-) converted to 2CO2
- (Also produces precursors for biosynthesis)
Lots of reducing power removes
What is catabolism stage 4
- Mitochondrial
- Electron transport and ATP synthesis
- NADH & FAD2H re-oxidised
- O2 required (reduced to H2O)
- Large amounts of energy substrate (ATP) produced
What is the general formula for carbohydrates?
(CH2O)n
Contain aldose or ketone groups
What are mono-, di-, oligo- and polysaccharides?
Mono: 3-9
Di: 2
Oligo: 3-12
Poly: 10-1000s
What are the 3 main dietary carbohydrates?
Glucose fructose galactose
Which cells have an absoute requirement of glucose
All tissues can metabolise glucose but some cells have an
absolute requirement.
• Red blood cells
• Neutrophils • Innermost cells of kidney medulla
• Lens of the eye (together approx. 40g/24 hours)
• Uptake depends on blood [glucose]
• CNS (brain) prefers glucose as fuel (approx. 140g/24 hours)
(can use ketone bodies for some energy requirements in times of starvation but needs time to adapt)
Need an inward gradient for absolute requires
What enzymes are present which digest carbohydrates
Saliva - Amylase - Starch, Glycogen -> dextrins
Note: lactose intolerance in some patients
Pancreas - Amylase -> monosaccharides
Small intestine - Disaccharidases attached to brush border membrane of epithelial cells • lactase (lactose) • sucrase (sucrose) • Pancreatic amylase ( a1-4 bonds) • isomaltase (a1-6 bonds)
Why is cellulose not digested in humans?
No enzymes to break down the B1-4 linkages present
Glycosidases only act on alpha
What is lactose intolerance
Primary lactase deficiency
• Absence of lactase persistence allele.
• Highest prevalence in Northwest Europe
• Only occurs in adults
Secondary lactase deficiency • Caused by injury to small intestine: • Gastroenteritis • Coeliac disease • Crohn's disease • Ulcerative colitis • Occurs in both infants & adults • Generally reversible
Congenital lactase deficiency
• Extremely rare, autosomal recessive defect in lactase gene. Cannot digest breast milk.
How are monosaccharides transported?
- Active transport (low to high concentration) into intestinal epithelial cells by sodium‐dependent glucose transporter 1 (SGLT1) then,
- passive transport (high to low concentration) via GLUT2 into blood supply
- Transport, via blood supply, to target tissues.
- Glucose uptake into target cells via facilitated diffusion using transport proteins (GLUT1 - GLUT5) (high to low concentration)
- GLUTs have different tissue distribution and affinities
What are glucose transporters
• Glucose uptake into cells from blood is via facilitated
diffusion using transport proteins (GLUT1 - GLUT5).
• GLUTs can be hormonally regulated (e.g. insulin/GLUT4)
GLUT1 : Fetal tissues, adult erythrocytes, blood–brain barrier.
GLUT2: Kidney, liver, pancreatic beta cells, small intestine
GLUT3 : Neurons, placenta.
GLUT4: Adipose tissue, striated muscle *insulin-regulated
GLUT5 : Spermatazoa, intestine
- GLUT2 and GLUT4 are particularly important and will be covered further elsewhere in your course.
WHAT ARE THE FUNCTIONS OF GLYCOLYSIS
Functions • Oxidation of glucose • NADH production (2 per glucose) • Synthesis of ATP from ADP (net= 2 ATP per glucose) • Produces C6 and C3 intermediates
WHAT ARE THE FEATURES OF GLYCOLYSIS
Features • Central pathway of CHO catabolism • Occurs in all tissues (cytosolic) • Exergonic, oxidative • C6 -> 2C3 (No loss of CO2) • With one additional enzyme (PDH), is the only pathway that can operate anaerobically • Irreversible pathway
What are 3 key enzymes of glycolysis
- Hexokinase (glucokinase in liver) - glucose -> glucose-6-phosphate
- Phosphofructokinase-1 - KEY CONTROL ENZYME - fructose-6-p -> fructose-1,6-bisphosphate
- Pyruvate kinase - phosphoenolpyruvate -> pyruvate
Why so many steps/enzymes?
- Chemistry easier in small stages
- Efficient energy conservation
- Gives versatility
• allows interconnections with other pathways
• allows production of useful intermediates
• allows part to be used in reverse - Allows for fine control
What is glycerol phosphate?
Important intermediate in glycolysis
• Important to triglyceride and phospholipid biosynthesis
• Produced from dihydroxyacetone phosphate (DHAP) in adipose tissue and liver
• Therefore, lipid synthesis in liver requires glycolysis
• (Note: Liver can phosphorylate glycerol directly)
What is 1,3-bisphosphoglycerate
Important intermediate in glycolysis
• Produced from 1,3-bisphosphoglycerate in RBC
• Important regulator of O2 affinity of haemoglobin (tense form)
• Present in red blood cells (RBC) at the same molar
concentration as haemoglobin (approx. 5 mM)
What is allosteric regulation of enzymes?
• Allostery (allo = other, steric = site) - activator/inhibitor binds at ‘another’ site
Proteins (usually enzymes) with two sites
1. Catalytic site
2. Regulatory site(s)
– Binding of specific regulatory molecule
– Affects catalytic activity
– can produce activation or inhibition
• Covalent modification (phosphorylation/dephosphorylation)
What are the principles of metabolic pathway regulation?
• Flux through pathways regulated in response
to the need
• Irreversible steps are potential sites of
regulation
• Reduced activity reduces the flux of substrates through the pathway directly
• Reducing levels of product
• Reversible steps are not regulated
• Reactions still come to equilibrium so leveksof product are unaffected
What is PFK?
Key regulator or glycolysis
Allosteric regulation (muscle)
• Inhibited by high ATP
• Stimulated by high AMP
Hormonal regulation (liver)
• Stimulated by insulin
• Inhibited by glucagon
Glucagon switches off enzyme - retain glucose in circulation - phosphorylates - inhibits
Insulin signalling high energy - use glucose - dephosphorylates - stimulates
How else is glycolysis regulated
Metabolic regulation Product inhibition by g6-p High [NADH] or low [NAD+] = high energy level signal • Causes product inhibition of step 6 • Inhibits glycolysis
Hormonal activation • PFK and Pyruvate kinase • increase by high insulin:glucagon ratio • i.e. high insulin low glucagon • (enzyme dephosphorylation)
Hexokinase product inhibition by g6-p
Describe step 6
• 2 moles of NADH produced per mole of glucose
• Pathway therefore requires NAD+
• Total NAD+ + NADH in cell is constant, therefore, glycolysis would stop when all NAD+ is converted to NADH
• Normally, NAD+ is regenerated from NADH in stage 4 of metabolism (see Lecture 4) BUT
1. RBC have no stage 3 or 4 of metabolism
2. Stage 4 needs O2 - supply of O2 to muscles and gut often reduced
• Therefore, need to regenerate NAD+ by some
other route
Lactate Dehydrogenase (LDH)
How is lactate produced
Produced from glucose (and alanine) via pyruvate
• without major exercise 40 - 50 g/24 hours
– RBC, skin, brain, skeletal muscle, G.I. tract
• Strenuous exercise (includes hearty eating!) 30 g/5
min
– Plasma levels x 10 in 2 - 5 min
– Back to normal by 90 min
• Pathological situations, e.g.
– SHOCK
– CONGESTIVE HEART DISEASE
What is the lactate dehydrogenase reaction?
NADH + H+ + pyruvate NAD+ + lactate
CH3COCOOH -> CH3CHOHCOOH
Catalysed by LDH
• Lactate by liver and heart (via LDH)
• produced by RBC and skeletal muscle (skin, brain, GI)
• Released into blood and
• normally metabolised by liver and heart
• Liver and heart need NAD+ to be regenerated efficiently, usually
well supplied with oxygen
Keeps glycolysis running
Where is LDH found?
RBC, skeletal muscle (skin, brain, GI)
In low O2 conditions pyruvate converted to lactate instead of stage 4 of metabolism which requires O2
Impaired in liver disease
Vitamin deficiency - thiamine
Alcohol NAD+ -> NADH
Enzyme deficiencies
What is plasma lactate concentration determined by?
Plasma concentration determined by relative rates of
- production
- utilisation (liver, heart, muscle)
- disposal (kidney)
What ate hyperlactaemia and lactate acidosis?
Hyperlactaemia
• 2 - 5 mM
• Below renal threshold
• No change in blood pH (buffering capacity)
Lactic acidosis
• above 5 mM
• Above renal threshold
• Blood pH lowered
Describe fructose metabolism and its clinical importance
Fructose = fruit monosaccharide
Glucose-fructose disaccharide
Metabolised in liver in human
Clinical Importance 1. Essential fructosuria - fructokinase missing • Fructose in urine no clinical signs 2. Fructose intolerance - aldolase missing • Fructose-1-P accumulates in liver - leads to liver damage • Treatment? - remove fructose from diet
Describe galactose metabolism
Lactose, dissacharide in milk, galactose + glucose Metabolism in liver (major tissue)
UDP-glucose acts catalytically Clinical importance - Galactosaemia - 1 in 30,000 births
What is galactosaemia
Autosomal recessive disorder which affects how the body processes galactose
Deficiency in any of the enzymes galactokinase, uridyl transferase or up-galactose epimerase can cause it
Milk rich diet - infancy
Unable to utilise galactose
Galactokinase deficiency (rare) - galactose accumulates
Transferase deficiency (common) - galactose and galactose-1-P accumulate Problem
• Galactose enters other pathways
Galactose —> galactitol
(NADPH -> NADP+)
Catalysed by aldose reductase
- depletes lens of NADPH - structure damaged - cataracts
- Accumulation of galactose-1-P affects liver, kidney, brain - How?
- Treatment - no lactose in diet
- Raised galactose concentration enters new pathways
- Depletes NADPH levels
- Prevents maintenance of free sulphydryl groups on proteins
- Inappropriate disulphide bond formation
- Loss of structural and functional integrity of some proteins that depend on free -SH groups
- E.g. lens of eye
What is the pentose phosphate pathway?
Series of non-oxidative reactions convert 5C sugars into 6C and 3C sugars which can be utilised by glycolysis
All enzymes in cytosol
2 stages
1) oxidative decarboxylation
Glucose-6-p —-> C5 sugar + CO2
(NADP+ —> NADPH)
2) rearrangement to glycolysis intermediates
3 C5 sugars ——> 2 f6P + 1 glyceralddehyde-3-p
No atp production
Loss of co2 to irreversible
Controlled by NADP+/NADPH ratio at G6P dehydrogenase
What are the functions of the pentose phosphate pathway?
- produce NADPH in cytoplasm
(a) Biosynthetic reducing power
e.g. lipid synthesis therefore, high activity in liver and adipose tissue
(b) Maintain free -SH (cysteine) groups on certain proteins
Prevent oxidation to - S - S - (disulphide bonds) - Produce C5 sugar for nucleotides needed for nucleic acid
synthesis
Therefore, high activity in dividing tissues e.g. bone marrow
Describe G6PDH deficiency
- Pentose phosphate pathway has an important role in providing NADPH to maintain SH group of proteins in a reduced state
- Structural integrity and, hence, functional activity of some proteins depends on free -SH groups
- G6PDH deficiency is a very common inherited defect
- e.g. in RBC, low NADPH -> disulphide bonds formed -> aggregated proteins, Heinz bodies -> haemolysis
- Lens of eye
What are the key points of the pentose phosphate pathway?
• Starts from Glucose-6-phosphate • Important source of NADPH required for: • Reducing power for biosynthesis • Maintenance of GSH levels • Detoxification reactions • Produces C5-sugar ribose required for synthesis of : • Nucleotides, • DNA & RNA. • No ATP synthesised. CO2 produced • Rate limiting enzyme is: Glucose 6-phosphate dehydrogenase LEARN PRINCIPLES ONLY: No need to memorise entire pathway!
What is pyruvate dehydrogenase?
• Mitochondrial matrix
(pyruvate transported from cytoplasm across mitochondrial membrane)
• PDH is a large multi-enzyme complex (5 enzymes)
• The different enzyme activities require various cofactors (FAD, thiamine pyrophosphate and lipoic acid). B-vitamins provide these factors, so reaction is sensitive to Vitamin B1 deficiency.
• Reaction is irreversible, so is a key regulatory step.
• Irreversible loss of CO2
• Pyruvate cannot be formed from acetyl-CoA
• Large multi-enzyme complex
• Irreversible reaction
• Subject to multiple regulation
• PDH deficiency – lactic acidosis
What is the TCA cycle?
- Mitochondrial
- A single pathway –
- Acetyl (CH3CO-) converted to 2CO2
- Oxidative (requires NAD+, FAD)
- Some energy produced (as ATP/GTP)
- (Also produces precursors for biosynthesis)
Give an overview of the tca cycle
Citrate (c6)
- > c6
- > isocritate (c6)
- > (nad+ to nadh, co2 released) c5
- > (coA in, nad+ to nadh, co2 released) c4
- > (gdp to gtp, coA out) c4
- > (fad to fadh2) c4
- > (H2O in) c4
- > (nad+ to nadh) c4
- > c4 + acetyl coA (c2) from link, makes citrate
How many tca cycles per glucose?
2
How is ca cycle regulated?
• Regulated by energy availability,
i.e. ATP/ADP ratio and NADPH/NAD+ ratio
Irreversible steps - release co2
Regulated by energy containing molecules
Adp is a low energy signal - stimulates
Enzymes: isocritate dehydrogenase, a-ketoglutarate dehydrogenase
Give a summary of the tca cycle
Mitochondrial
• Central pathway in the catabolism of sugars, fatty acids, ketone bodies, amino acids, alcohol
• Strategy - to produce molecules that readily lose CO2
• Breaks C-C bond in acetate (acetyl~CoA); carbons oxidised to CO2
• Oxidative producing NADH and FADH2
• Some energy as GTP (~= ATP) produced directly
• Produces precursors for biosynthesis
• Does not function in absence of O2 (c.f. electron transport chain)
• Intermediates act catalytically - no net synthesis or degradation of Krebs cycle intermediates alone
What happens in catabolism stage 4?
- Mitochondrial
- Electron transport and ATP synthesis
- NADH & FADH2 re-oxidised
- O2 required (reduced to H2O)
- Large amounts of energy (ATP) produced
How is reducing power used in aTp synthesis?
Two processes
1. Electrons on NADH and FAD2H transferred through a series of carrier molecules to oxygen (ELECTRON TRANSPORT)
Releases energy in steps
- Free energy released used to drive ATP synthesis
(OXIDATIVE PHOSPHORYLATION)
How are electrons transported
• Electrons are transferred through series of carrier molecules (mostly within proteins) to O2, with release of energy.
• ~30% of energy used to move H+ across membrane (a lot of the energy released as heat.)
• [H+] gradient (membrane potential) across inner mitochondrial membrane caused by energy of electrons = proton motive force
PTC = protein translocations complex
What is a Proton translocating ATPase?
ATP synthase
Uses hydrogen ion gradient to drive the synthesis of atp
Giving up energy of proton gradient
• Return of protons (H+) is favoured energetically by the
electrochemical potential (electrical and chemical gradient)
• Protons (H+) can only return across membrane via the ATP synthase and this drives ATP synthesis
What is oxidative phosphorylation?
• Electrons are transferred from NADH and FAD2H to
molecular oxygen
• Energy released is used to generate a proton gradient, proton motive force (pmf)
• Energy from the dissipation of the proton motive force is coupled to the synthesis of ATP from ADP
- Electrons in NADH have more energy than in FAD2H,
- so NADH uses 3 PTCs, FADH2 uses only 2.
- The greater the p.m.f. -> more ATP synthesized
- Oxidation of 2 moles of NADH -> synthesis of 5 moles of ATP (P/O = 2.5)
- Oxidation of 2 moles of FADH2 -> synthesis of 3 moles of ATP (P/O = 1.5)
How is oxyphos regulated?
• When [ATP] is high, i.e. [ADP] is low, so no substrate for ATP synthase (synthetase)
• so inward flow of H+ stops
• Concentration of H+ in the intermitochondrial space increases
• Prevents further H+ pumping - stops electron transport
• Normally oxidative phosphorylation and electron transport
are tightly coupled
• Both regulated by mitochondrial [ATP]
• High ATP = Low ADP
• When [ADP] is low, so no substrate for ATP synthase (synthetase)
• so inward flow of H+ stops
• Concentration of H+ in the intermitochondrial space increases
• Prevents further H+ pumping - stops electron transport
• Reverses with low [ATP]
How is oxidative phosphorylation inhibited
• Inhibitors block electron transport, e.g. cyanide (CN-) prevents acceptance of electrons by O2
Bind with higher affinity than oxygen - reduced h+ translocation reduces pmf - synthesis of ATP reduced - leads to cell death
No electron transport - Block the whole of oxyphos and all of catabolic
- Uncouplers increase the permeability of the mitochondrial inner membrane to protons - h+ enter mitochondria without driving ATP synthase
- dissipate the proton gradient, thereby reducing the proton motive force
- No drive for ATP synthesis
What are oxyphos diseases
Genetic defects in proteins encoded by mtDNA (some subunits of the PTCs and ATP synthase) so decrease in electron transport and ATP synthesis
=what is coupling of oxyphos
- What happens to the rest of the energy ?
- Lost as HEAT
- Efficiency depends on tightness of coupling
- Can be varied in some tissues
- Brown adipose tissue - degree of coupling controlled by fatty acids (uncouplers) - allows extra heat generation
Describe e- transport in brown adipose tissue
Contains thermogenin (UCP1) - naturally-occurring uncoupling
In response to cold, noradrenaline (norepinephrine) activates :
1. Lipase which releases fatty acids from Triacylglycerol
2. Fatty acid oxidation -> NADH/FADH2 -> electron transport
3. Fatty acids activate UCP1
4. UCP1 transports H+ back into mitochondria
So, Electron Transport uncoupled from ATP Synthesis. Energy of p.m.f. is then released as extra heat.
Family of UCPs – role in heat generation by uncoupling but may have other functions.
Where is brown adipose found
Newborn infants to mainatain heat around vital organs
Hibernating animals to generate heat to maintain body temp
Compare oxyphos and sub level phos
Oxy
Requires membrane-associated complexes (inner mitochondrial membrane)
Energy coupling occurs indirectly through generation & subsequent utilisation of a proton gradient (pmf)
Cannot occur in the absence of O2
Major process for ATP synthesis in cells requiring large amounts of energy
Sub
Requires soluble enzymes (cytoplasmic & mitochondrial matrix)
Energy coupling occurs directly through formation of high energy of hydrolysis bond (phosphoryl-group transfer)
Can occur to a limited extent in the absence of O2
Minor process for ATP synthesis in cells requiring large amouts of energy
