Exam 2 Flashcards
Galactokinase Deficiency
Elevated Galactose in the blood (galactosemia)and urine (galactosuria). Elevated galactisol = cataracts. Treatment is dietary restriction
Galactose 1 Phosphate uridylytransferase deficiency
Symptoms - Galactosemia, Galactosuria, vomiting and diarrhea
Effects- Liver damage , intellectual disability, cataracts
Treatment - Remove galactose from diet
Galactose Metabolism
From lactose in milk, digested by lactase in intestinal brush-borders.
*Lactose Intolerance from lactase deficiency causes diarrhea, bloating and
abdominal cramps following milk ingestion.
*Galactosemias are autosomal recessive diseases due to a defect in the gene
encoding galactokinase or galactose 1-uridyltransferase (classic galactosemia –
more severe).
*Patients have elevated levels of plasma galactose.
*Signs and symptoms of Classical Galactosemia include lethargy, vomiting after
ingesting lactose, hypotonia, mental retardation, cataracts and failure to thrive.
They are prone to bacterial infections eg
E.coli
*Jaundice (hyperbilirubinemia) results from the accumulated galactose 1-
phosphate which inhibits phosphoglucomutase (converts glucose 1-phosphate to
glucose 6-phosphate). This inhibits recycling of UDP-glucose that is used for
UDP-glucuronate synthesis - UDP-glucuronate is used for bilirubin conjugation.
*Treatment involves a galactose free diet.
*In well fed states glucose 1-phosphate is
channeled into glycogenesis.
*In hypoglycemia, galactose administration
can increase blood glucose levels.
*Galactose accumulation in the blood can lead
to its conversion to galactitol in the lens by
aldolase reductase causing osmotic swelling
and cataract formation in the lens and nerve
damage.
This is why
diabetics are
at risk.
These pathways occur in the
lens, kidneys and Schwann
cells. They lack sorbitol
dehydrogenase which
results in cataracts,
retinopathy and peripheral
neuropathy
Fructose Metabolism
From fruits, honey, sucrose(table sugar),corn syrup and
sorbitol. Most tissues phosphorylate fructose through
hexokinase slowly but the liver and kidneys use
fructokinase.
*Because the DHAP and glyceraldehyde 3-phosphate are
downstream from the key regulatory step in glycolysis
(PFK1), fructose is a quick source of energy and can also be
readily used for triglyceride biosynthesis.
Fructokinase deficiency (Essential Fructosuria)
mild disorder, Benign conditin, Auto Somal recessive , fructose not trapped in cells, no cataract (ketone) fructosuria. is an
autosomal recessive disorder with a mild phenotype
because fructose is not trapped within cells, there is no
cataract because fructose is a ketose not an aldose.
Aldolase B deficiency
Fructose 1 accumulation = depletion of ATP
Inhibits production of dihydroxyacetone and glyceraldehyde to produce pyruvate = no glucose made
Symptoms : hypoglycemia , vomiting, jaundice , hemorrhage, hepatomegaly , renal dysfunction , hyperuricemia
Primary Purposes for Pentose Phosphate Pathway
Biosynthetic:
NADPH
Ribose 5-phosphate and others
Protective:
NADPH
* Metabolism of
xenobiotics
* Removal of reactive
oxygen
Can operate in several different
modes depending on cellular
conditions
Two branches: Oxidative and non-
oxidative
Pentose Phosphate Pathway key facts
NADH - has a phosphate group to allow certain molecule to bind like nucleotides
An alternate pathway of glucose metabolism
Occurs in the cytoplasm of all cells especially
lactating mammary glands, liver, adrenal cortex
and RBCs
Does not yield ATP
It leads to the formation of NADPH for fatty acid
biosynthesis and maintaining reduced glutathione
for antioxidant activity
It also leads to the formation of ribose sugars for
nucleic acid synthesis.
Pentose Phosphate Pathway main functions
Generate NADPH
Pentose Sugar Ribose
Pentose Phosphate Pathway happens when
NADPH is low
Insulin triggered
Oxidative
Stops at Ribose 5 Phosphate
Produced 2 NADPH
DNA formation
Non Oxidative
Bring metabolites back into glycolysis to gluconeogenesis pathway to form intermediates
Pentose Pathway Location
Cytoplasm of all cells especially lactating mammary glands , liver adrenal cortex and RBC
Key Steps in the pathway
The rate-limiting step is catalyzed by Glucose 6-phosphate
dehydrogenase(G6PD)
It involves an irreversible oxidative phase involving G6PD and 6-
phosphogluconate dehydrogenase with the production of ribulose 5-
phosphate and the production of NADPH.
G6PD is induced by insulin and inhibited by NADPH and activated by NADP
This is followed by the reversible oxidative phase beginning with ribulose 5-
phosphate producing an equilibrated pool of sugars used in biosynthetic
reactions
Fructose 6-phosphate and glyceraldehyde 3-phosphate are intermediates
of this pathway and can be channeled into glycolysis directly.
Transketolase is important for these interconversions and is thiamine-
dependent. It is the only thiamine dependent enzyme in erythrocytes.
(can be used to evaluate a patient’s nutrituional status for thiamine
USES OF
NADPH
- Relax smooth muscles
- Prevent platelet aggregation
- Acts as a neurotransmitter in
brain - Mediates tumoricidal and
bactericidal actions of
macrophages
G6PD Deficiency
X-linked recessive disease, common in areas where malaria is
endemic.
Peroxides generated in RBCs are destroyed by glutathione
peroxidase/reductase system (which uses NADPH. Other cells
that produce NADPH by using alternate enzymes like malate
dehydrogenase are not susceptible to this oxidative damage
like RBCs are.
Patients have episodic acute hemolysis because of
accumulated ROS, protein denaturation and lipid peroxidation
– membrane fragility.
Fava beans(favism), Infections, moth balls (naphthalene) and
some drugs(primaquine, dapsone, isoniazid) predispose to
hemolysis in these patients – strong oxidizing agents!
Carbohydrate
Catabolism
Key Point:
* The citric acid cycle IS the Engine that drives energy
production regardless of the source:
* Carbohydrate -> acetyl CoA
* Lipid -> acetyl CoA
* Protein -> acetyl CoA
* Certain nucleotides -> acetyl CoA
How does NADH
get into the
mitochondria?
Very polar: ie cannot diffuse
through the mitochondrial
membrane.
- NADH doesn’t actually cross the
membrane: The electrons are
moved across the membrane:
Organ specific transport
mechanisms:
Heart and Liver use a common transport
mechanism
* Brain and skeletal muscle use a different
mechanism
Transport of NADH into the mitochondria
TCA occurs in the mitochondria: NADH is already inside
Glycolysis occurs in cytosol: NADH must be
transported
Complex I
NADH + Q-> NAD+
+ QH2
* Electrons enter from the matrix
* H+ ions are “pumped” out of the mitochondria in
response to the electrons moving through the
complex from NADH to Q
* This transfer is highly favorable
* One arm extends
into the matrix
* The other is fully
membrane
bound
* CoQ: Eo’ = 0.045
Complex II
Succinate + Q -> Fumarate
+ QH2
Complex II
* Not a pump, so no contribution to the membrane
potential or ATP formation
* Like complex I, electrons are delivered to Q
* Both Complex I and II contribute to the pool of QH2
in the mitochondrial membrane
Complex III
QH2 + 2Cytc(ox) -> Q + 2H+ + 2Cytc(red)
DEo’ = 0.19 V
DGo’ = -36.6 KJ/mol
contains high and low potential heme cofactors
- Stoichiometry changes: 1 QH2 reduces 2
cytochrome c - Cytochrome c is a peripheral membrane protein
that migrates along the out surface of the inner
mitochondrial membrane.
Complex III is also a “pump” that moves H+ ions
across the membrane from inside to outside and
helps to generate the membrane potential.
Complex IV
4 Cytcred + O2 + 4
H+–> 4 Cytcox + 2 H2O
- Electrons finally reach their ultimate home.
- Lowest energy state for the electrons is on oxygen
in the form of water. - The movement of these electrons to their lowest
energy state is coupled to the generation of a
membrane potential (a charged battery) - The mitochondrial membrane is now ready to
produce ATP by allowing a H+ “current” to pass
through.
How does electron flow drive H+ translocation and what is it’s significance?
Energy is stored in the proton gradient!
ATP Synthase: Features
Lolipop structure
F1 subunit extends into the matrix of the mitochondria
This is where ATP is synthesized
F0 unit is membrane bound
Molecular Motion in ATPase
A look down the 3-fold axis of the F1 ATPase.
Channel is lined with hydrophobic amino acids.
Perfect for slippery surface for rotation
Why is rotation important?
Binding change mechanism
ATP release is key to catalysis
In T-state, ATP synthesis occurs spontaneously
Transport of Material
ATP has -4 charge
ADP has -3 charge
Proposed Rotary Action
Protons want to move from bottom to top.
Only way to move is through rotation of the c ring with respect to the alpha3beta3
Uncoupling: A source of Heat
Hibernating animals can generate heat this way. “Brown Fat”
Honey bees and futile cycles
HYPOPHOSPHATEMIA AND GLYCOLYSIS
May be caused by parenteral nutrition, malnourishment(refeeding syndrome), chronic alcohol abuse, diabetic ketoacidosis, sepsis, respiratory alkalosis, or primary hyperparathyroidism, Fructosemia and galactosemia
In glycolysis, it inhibits;
Glyceraldehyde 3-phosphate dehydrogenase
Impairs mitochondrial ATP synthesis
Diminishes 2,3BPG levels and impairs tissue oxygenation
Increased hemolysis and impaired leukocyte function (sepsis)
Nucleotide Kinase
: Forms UTP from UDP
Myokinase
Creates ATP from 2 ADP: Also form AMP which is a key regulator , and also involved in the AMP cycle to boost TCA activity
Creatine phosphate
High energy phosphate “buffer” in muscle cells to regenerate ATP immediately after exercise begins.
Insulin Secretion
Insulin secretion in beta cells is triggered by rising blood glucose levels.
Glucose uptake by the GLUT2 transporter leading to cell depolarization, calcium influx leading to the exocytotic release of insulin from their storage granule.
3-Steps in Glycogen breakdown
I. release of glucose 1-phosphate by the phosphorylase (PLP-dependent) Vitamin B6
II. phosphoglucomutase
III. debranching (transglycosylase and hydrolysis).
Glycogen Breakdown
Glycogen Phosphorylase
Pyridoxal phosphate (B6) dependent
Phosphorolysis (ie like hydrolysis)
Energy conservation in the phosphate linkage.
Produces glucose 1-phosphate
Key enzyme in regulating breakdown is glycogen phosphorylase
Hormonal: Insulin and glucagon
AMP activation
G1P and G6P inhibition
Regulation is tightly coordinated with the activity of the Glycogen Synthase, which regulates synthesis.
Glycogen Synthesis
Different mechanism for synthesis than for breakdown.
This was realized when the cause of McArdle’s disease was discovered.
Defective phosphorylase
Energy requirement:
UTP used as an energy source
DG under physiological conditions is +5 to +8 KJ/mol.
Introduction to “energy coupling”
Three activities required for synthesis from G1P
I. UDP-Glucose Pyrophosphorylase
II. Synthase
III. Branching enzyme
UDP-glucose Pyrophosphorylase
Release of pyrophosphate for subsequent hydrolysis provides additional thermodynamic driving force.
Branching Enzyme
7 residue fragment is transferred from one branch to start another
Transferred branch must come from a chain of at least 11
Regulation of Glycogen metabolism
Synthesis and breakdown are both spontaneous under normal cellular conditions. (Just like glycolysis and gluconeogenesis)
Requires tight regulation
Phosphorylase and Synthase are inversely regulated by the same cellular signal
Hormonal regulation of Liver phosphorylase:
Key hormones: glucagon, epinephrine, Insulin
Diabetes Melitus : Type I
: Autoimune disease resulting from destruction of pancreatic b-cells responsible for insulin production.
Sometimes called insulin sensitive, because administration of insulin will restore blood glucose level.
Diabetes Melitus : Type II
Adult onset: results from a continuing desensitization of cells toward insulin. Cannot be treated with insulin.
Diabetes Melitus
Both cause hyperglycemic episodes, which can damage blood vessels and create a host of other medical conditions (glaucoma).
underlying causes of diabetes melitus
Glut 1 moves glucose across to fetus
A hyperglycemic mother will lead to hyperglycemic fetus
Endocrine system of the fetus controls how the glucose is utilized
Normal fetus has anabolic conditions (synthesis and storage)
Hyperglycemic fetus produces more insulin
Glut 4 is produced in late gestational stage in muscle and fat.
This explains the large size of infants of diabetic mothers
Normal transitioning after child-birth
At birth: Glucose supply is cut off
Approximately 7 min worth of glucose
Initial depletion of glucose results in mobilization of liver glycogen
This results from hormonal changes
Beyond 12 hours, gluconeogenesis is the primary source of glucose from amino acids that are glucogenic
Fatty acids also provide energy to compensate as glucose falls: glucagon stimulate lipolysis and release of fatty acids Brain will switch to ketone bodies, heart turns to fatty acid
Diabetic Problem
Endocrine environment of the newborn from diabetic mother is poised to remove glucose:
Very high insulin: Double that of normal infant.
Effects of Insulin: Summary
Inhibits ketogenesis
Inhibits lipolysis
Inhibits gluconeogenesis
Inhibits glycogen breakdown
Stimulates lipid synthesis
Stimulates glycogen synthesis
Stimulate glycolysis
Pyruvate Carboxylase Deficiency
Symptoms range from benign to severe and include fasting hypoglycemia, hypothermia, hypotonia, neurologic dysfunction, vomiting, lactic/ketoacidosis.
Pyruvate carboxylase catalyzes the formation of oxaloacetate from pyruvate and CO2 in the mitochondrial matrix.
It’s biochemical significance involves gluconeogenesis, anaplerosis and lipogenesis.
It is exported by the malate aspartate shuttle into the cytoplasm for conversion to phosphoenolpyruvate unto glucose
Oxaloacetate is used for the generation of intermediates for the TCA cycle
Aspartate is formed by transamination of oxaloacetate in the mitochondrial matrix but in PC deficiency, aspartate levels will be low – impaired gluconeogenesis and electron transfer for ATP generation.
An impaired electron transfer lowers the redox equilibrium in the mitochondrial matrix lowers the β-hydroxybutyrate/acetoacetate ratio - ketoacidosis
The cytoplasmic redox equilibrium promotes the formation of lactate from pyruvate leading to lactic acidosis and reducing the supply of acetyl-CoA to the TCA cycle.
Low aspartate levels lead to hypercitrullinemia, hyperammonemia and low plasma glutamate levels.
PC deficiency impairs lipogenesis due to the depletion of pyruvate, oxaloacetate and acetyl-CoA required for fatty acid biosynthesis – widespread demyelination of cerebral and cerebellar white matter characteristic of the disease.
Alcohol and Hypoglycemia
In addition to poor nutrition, alcohol metabolism to acetyl-CoA generates a lot of NADH in the cytoplasm
NADH favors the formation of lactate from pyruvate, malate from OAA and glycerol 3-phosphate from DHAP
The large amounts of NADH inhibit gluconeogenesis and favor lipid accumulation in the liver and excessive storage of triglycerides(alcoholic steatosis). They also delay fatty acid oxidation
So with exercise and accumulation of lactate, NAD+ is not available for conversion of lactate to pyruvate because it is used to metabolize alcohol.
All these promote hypoglycemia!
Lingual lipases
Act on TAGS in the saliva and into the stomach.
Gastric Lipases
small amounts active at low pH
enterogastrone hormones secreted by the duodenal mucosa:
inhibits HCl production and forward movement of gastric contents (ie, slows gastric emptying) allowing the churning action of the stomach
Intestinal lipases
Small intestines
Steapsin (a lipase) and cholic acids (other bile salts)
Secreted by Pancreatic and Bile ducts
Uptake and /or storage of fats
In adipose tissue, liver, muscle etc….
Lipoprotein Lipases in the blood vessel walls release FAs and glycerol (attached to vessel walls by Heparin-sulfate)
Apolipo CII binds LPL, which activates the lipoprotein lipases on vessel walls
FAs are taken up by the tissue.
Glycerol is generated from glucose and TAGs are synthesized
Steps in Fatty acid Metabolism
Hormone sensitive lipases release Free Fatty acids from adipose tissue
Free fatty acids travel in the blood associated with plasma proteins (Albumin)
Uptake by peripheral tissue is passive, thus dependent on concentration
Free diffusion
Fatty acid transport proteins
Breakdown of fatty acids for fuel.
FAs are transported via a FA transport protein (along with Na+ into the cell).
FAs are taken up in the cell and transported (chauffeured) by FA binding proteins.
They are activated via attachment to CoA-SH
Transported inside the mitochondria via Carnitine transporter and acyl transferase system
b-oxidation cycle generates acetyl-CoA:
No need to know a- or w-oxidation
Fat metabolism includes the following processes
- Digestion, absorption, and transport of dietary fat
- Generation of metabolic energy from this fat
- Storage of excess fat in adipose tissue
- Significant Metabolic links between triglycerides and other biomolecules, including carbohydrates and ketone bodies
FATTY ACIDS: Structure review
Fatty acids are long-chain carboxylic acids (R-COOH)
The carboxyl carbon is number 1 and carbon number 2 is the α carbon.
When naming fatty acids, the number of carbon atoms and double bonds are used together eg palmitic acid (C16:0), Linoleic acid (C18:2)
Saturated fatty acids do not have double bonds while unsaturated fatty acids have double bonds.
Saturated fatty acids in plasma membranes restrict membrane fluidity unlike unsaturated fatty acids
Defects related to FA metabolism
Systemic Carnitine Deficiency
Zellweger’s syndrome (spectrum Disorder)
X-linked adrenoleukodytrophy
C-P transferase deficiency (Type I and II)
Jamaican Vomiting Sickness: (ACDH) Hypoglycin A
SIDS (MCACDH)
Fatty acid – glucose relationship
Glucose can be converted to Fatty Acids via production of acetyl CoA through Glycolysis and PDH
Fatty Acids cannot be converted to glucose because the PDH reaction is irreversible and there is no other metabolic route in humans.
Exception: Odd chain Fatty acids: BECAUSE
Synthesis of fatty acids from Acetyl CoA
Occurs in the cytosol
mitochondrial conditions influence synthesis
Fatty Acid Regulation
NADH, ATP: internal
Insulin: external
High ATP -> low ETC -> High NADH -> Low TCA -> High Citrate
Citrate shuttle and Malic enzyme
The citrate shuttle transports acetyl-CoA groups (as citrate) from the mitochondria to cytoplasm for fatty acid synthesis. This is promoted by high NADH, ATP and insulin.
In the cytoplasm, ATP-citrate lyase splits citrate into acetyl-CoA and oxaloacetate. The oxaloacetate is returned to the mitochondria: malic enzyme supports this by converting malate into pyruvate This generates NADPH supplementing HMP shunt (PPP).
Acetyl-CoA carboxylase activates acetyl-CoA and catalyzes the rate-limiting step in fatty acid biosynthesis. It requires biotin, ATP and CO2. Its controls include insulin and citrate activation.
The CO2 in not incorporated into the fatty acid but removed by the subsequent step (fatty acid synthase)
Acetyl-CoA Carboxylase
Acetyl-CoA Carboxylase catalyzes the rate-limiting step in FA synthesis
Sources of acetyl-CoA include pyruvate, isoleucine, lysine, leucine, tryptophan (? High protein diet)
It uses biotin as a co-factor.
The ATP-dependent carboxylation of the biotin is followed by a transfer of the carboxyl group to acetyl-CoA to form malonyl-CoA.
It is induced by insulin (dephosphorylation), citrate and ATP but inhibited by Palmitoy-CoA, AMP, glucagon and epinephrine
The summary is:
HCO3- + ATP + acetyl-CoA ADP + Pi + malonyl-CoA
METFORMIN
Oral hypoglycemic agent used in Insulin-independent diabetes mellitus and insulin-resistance (Polycystic Ovarian Syndrome)
Reduces blood glucose levels and promotes weight loss
Inhibits Complex I of the ETC leading to increased AMP/ATP ratio
Increased AMP inhibits F1,6 bisphosphatase and activates PFK-1
AMP activates AMPK which increases GLUT4 expression, It also inhibits triglyceride synthesis and lipolysis
Acetyl-CoA Carboxylase:
Acetyl-CoA Carboxylase catalyzes the rate-limiting step in FA synthesis
Sources of acetyl-CoA include pyruvate, isoleucine, lysine, leucine, tryptophan (? High protein diet)
It uses biotin as a co-factor.
The ATP-dependent carboxylation of the biotin is followed by a transfer of the carboxyl group to acetyl-CoA to form malonyl-CoA.
It is induced by insulin (dephosphorylation), citrate and ATP but inhibited by Palmitoy-CoA, AMP, glucagon and epinephrine
The summary is:
HCO3- + ATP + acetyl-CoA -> ADP + Pi + malonyl-CoA
Fatty acid Synthase (FAS)
Also called palmitate synthase because palmitate is the first fatty acid synthesized de novo in humans.
It contains an acyl carrier protein(ACP) that requires pantothenic acid. It has 2 sulfhydryl groups at Pantothenate and cysteine
Malonyl-CoA is the substrate/primer for FAS, but only the carbon from acetyl-CoA is incorporated into the fatty acid.
Acetyl-CoA is used as a primer then malonyl-CoA is used to add subsequent C2 to form even-numbered fatty acids while propionyl-CoA is used to form odd-chain fatty acids(in ruminant fats and milk)
NADPH is the electron donor and is used to reduce the acetyl groups added to fatty acid (from PPP and malic enzyme)
Eight acetyl-CoA groups are required to produce palmitate(16:0) (7 converted to Malonyl and 1 acetyl)
The sequence of elongation reactions is repeated until a saturated 16-carbon acyl CoA has been assembled.
Finally:
When the fatty acid is 16 carbon atoms long, a Thioesterase domain catalyzes hydrolysis of the thioester linking the fatty acid to phosphopantetheine.
The 16C saturated fatty acid palmitate is the final product of the Fatty Acid Synthase complex.
fatty acid summary
The equation for synthesis of palmitate from acetyl-CoA, listing net inputs and outputs:
8 acetyl-CoA + 14 NADPH + 7 ATP
palmitate + 14 NADP+ + 8 CoA + 7 ADP + 7 Pi
Summary based on malonate as an input:
acetyl-CoA + 7 malonyl-CoA + 14 NADPH
palmitate + 7 CO2 + 14 NADP+ + 8 CoA
Fatty acid synthesis occurs in the cytosol. Acetyl-CoA generated in mitochondria is transported to the cytosol via a shuttle mechanism involving citrate.
Elongation beyond the 16-C length of the palmitate product of Fatty Acid Synthase is mainly catalyzed by enzymes associated with the
endoplasmic reticulum (ER).
ER enzymes lengthen fatty acids produced by Fatty Acyl Synthase as well as dietary polyunsaturated fatty acids.
Fatty acids esterified to
coenzyme A serve as substrates.
is the donor of 2-carbon units in a reaction sequence similar to that of Fatty Acid Synthase except that individual steps are catalyzed by separate proteins.
Malonyl-CoA
introduce double bonds at specific positions in a fatty acid chain.
Desaturases
Mammalian cells
are unable to produce double bonds at certain locations, e.g., ∆12
some polyunsaturated fatty acids are dietary essentials, e.g.,
linoleic acid, 18:2 cis ∆9,12 (18 C atoms long, with cis double bonds at carbons 9-10 & 12-13).
Formation of a double bond in a fatty acid involves the following endoplasmic reticulum membrane proteins in mammalian cells:
NADH-cyt b5 Reductase, a flavoprotein with FAD as prosthetic group.
Cytochrome b5, which may be a separate protein or a domain at one end of the desaturase.
Desaturase, with an active site that contains two iron atoms complexed by histidine residues.
The desaturase catalyzes a mixed function oxidation reaction.
There is a 4-electron reduction of O2 2 H2O as a fatty acid is oxidized to form a double bond.
2e- pass from NADH to the desaturase via the FAD-containing reductase & cytochrome b5, the order of electron transfer being:
NADH FAD cyt b5 desaturase
2e- are extracted from the fatty acid as the double bond is formed.
E.g., the overall reaction for desaturation of stearate (18:0) to form oleate (18:1 cis D9) is:
stearate + NADH + H+ + O2 oleate + NAD+ + 2H2O
