The Citric acid cycle (2)

Report
Dr.S.Chakravarty M.D.
Carbohydrates
Fats
recycling
Proteins
3 Stages Of
Metabolism
1
2
3
The citric acid cycle (Krebs cycle, tricarboxylic acid
cycle) is a sequence of reactions in mitochondria that
oxidizes the acetyl moiety of acetyl-CoA and reduces
coenzymes that are reoxidized through the electron
transport chain, linked to the formation of ATP.
How does Pyruvate enter mitochondria?
• Symport along with H+ ions
The Enzyme subunits
• E1 - Thiamine pyro phosphate (TPP) (B1)
• E2 – Lipoic acid
Co-enzyme-A – (Pantothenic acid)
• E3 – NAD – Niacin (B3)
FAD – Riboflavin (B2)
Tender – Thiamine
Loving - Lipoamide
Care -CoASH
For – (FAD)Riboflavin
Nancy- NAD (Niacin)
3 Enzymes:1)PDH,
2)α-KGDH(TCA cycle)
3)Branched keto acid
dehydrogenase
Cofactor
Location
Function
TPP(Thiamine
pyrophosphate)
Bound to E1
Decarboxylates pyruvate
yielding a hydroxyethyl TPP
carbanion
Lipoic acid
Covalently linked to a Lys
on E2(lipoamide)
Accepts the hydroxyethyl
carbanion from TPP as an
acetyl group
Co A( Coenzyme A)
Substrate for E2
Accepts acetyl group from
lipoamide
FAD(Flavin adenine
dinucleotide)
Bound to EE3
Reduced by lipoamide
NAD+
Substrate for E3
Reduced by FADH2
• PDH is IRREVERSIBLE ( Fats cannot be converted to
glucose.)
• COMMITTED STEP in oxidation of glucose.
• ENERGETICS :- 1 NADH IS GENERATED = 2.5 ATP
• REGULATION :– End product as well as covalent modification
– Phosphorylation of enzyme by a kinase decreases the
activity and dephosphorylation decreases the activity.
Regulation of PDH enzyme:
1. Regulation by end
product inhibition
(Allosteric)
2. Regulation by Covalent modification:
• PDH kinase –
inactivation of
enzyme
• PDH Phosphatase activation
Congenital Lactic acidosis:
• Deficiency of Pyruvate Dehydrogenase enzyme.
• Inability to convert Pyruvate to Acetyl co-A.
• Shunted to Lactate Dehydrogenase to form Lactic
Acid.
• Deficient NADH leading to deficient ATP
• Lactic acidosis, severe psychomotor retardation,
damage to brain stem, cortex etc.
Other causes of lactic acidosis:
Reasons:
Type:
• Severe exercise
•
•
Mercury poisoning
Arsenic poisoning
•
Pyruvate carboxylase deficiency
•
TPP deficiency
•
Chronic Alcoholism
excess lactate
•
Binds to –SH groups of Lipoic acid and
forms a stable complex.
• Decreased absorption and
poor diet.
• The citric acid cycle is the final common pathway for the
oxidation of carbohydrate, lipid, and protein because glucose,
fatty acids, and most amino acids are metabolized to acetylCoA or intermediates of the cycle.
• It is a source of reduced co-enzymes that provide the
substrates for the respiratory chain.
• It is both catabolic and anabolic (amphibolic).
contd..
• It also has a central role in gluconeogenesis, lipogenesis, and
interconversion of amino acids.
– So, components of the cycle have a direct or indirect controlling effects
in key enzymes of other pathways.
• Many of these processes occur in most tissues, but the liver is the
only tissue in which all occur to a significant extent.
– The repercussions are therefore profound when, for example, large
numbers of hepatic cells are damaged as in acute hepatitis or replaced
by connective tissue (as in cirrhosis).
• Very few, if any, genetic abnormalities of citric acid cycle enzymes
have been reported; such abnormalities would be incompatible
with life or normal development.
• Tissues :- All tissues
• Subcellular site :- Mitochondrial Matrix
NADH
Enzyme
bound
FADH2
ATP
Substrate level
phosphorylation
NADH
NADH
Thiamin, lipoate , FAD
Enzyme
bound
- THE SODA POP and
the world’s food Supply ?
• Cirtate – fruity flavour – used commercially in soft drinks .
• PLASTICIZER and FOAM INHIBITOR
• INDUSTRIALLY PRODUCED using fungus Aspergillus Niger
• ALUMINIUM(Al +3) ions –MOST ABUNDANT
METAL IN EARTH’S CRUST-extremely toxic to
PLANTS
PLANTS SECRETE CITRATE INTO THE SOIL WHICH CLELATES Al+3
Genetically engineered plants which secrete 5-6 times normal levels of citrate
in soil 
OAA is viewed as a catalyst , which enters into the cycle , causes complete
oxidation of acetyl CoA , and is regenerated in the end without any loss.
Reaction catalyzed by
Method of production
ATP molecules formed
Isocitrate dehydrogenase
Respiratory chain oxidation
of NADH
2.5 (3)
-ketoglutarate
dehydrogenase
Respiratory chain oxidation
of NADH
2.5 (3)
Succinate thiokinase
Substrate level
phosphorylation
Succinate dehydrogenase
Respiratory chain oxidation
of FADH2
1.5(2)
Malate dehydrogenase
Respiratory chain oxidation
of NADH
2.5 (3)
1
Net –>
10 (12)
1)Final common oxidative pathway
2)Fat is burnt on the wick of carbohydrates
• Oxidation of fats need the help of Oxaloacetate
which enters into the cycle and is regenerated in the
end .
• The major source of OAA is Pyruvate. (Carbohydrate)
3) Excess carbohydrates are converted to neutral fats via citrate and
ATP-citrate lyase but not vice versa because Pyruvate dehydrogenase
step is irreversible.
P
IRREVERSIBLE
T
4)Amphibolic ( Catabolic and Anabolic )
FATTY
ACIDS,
STEROLS
gluconeogenesis
HEME
GABA
5) TCA cycle plays an important role in Gluconeogenesis ,
Transmination and Deamination.
6) Anaplerotic ( filling – up) reactions
-> As shown before ,TCA cycle acts as precursors of biosynthetic
pathways , e.g Heme .
So, there is constant efflux of carbon units from the cycle .To
counterbalance the loss , filling up reactions are necessary .
Eg. - Pyruvate to Oxaloacetate(PYRUVATE CARBOXYLASE) ( most
important)
-- Phosphoenolpyruvate to Oxaloacetate (PEP CARBOXYLASE)
--Pyruvate to Malate (Malic enzyme)
7) Metabolic traffic regulator
-All metabolisms end in TCA.
-Availability or lack of intermediates govern the directions of
pathways converging or going out of TCA.
• Regulation of the Citric Acid Cycle Depends Primarily on a Supply of
Oxidized Cofactors.
• Individual enzymes of the cycle are regulated - The most likely sites
for regulation are the nonequilibrium reactions catalyzed by
pyruvate dehydrogenase, citrate synthase, isocitrate
dehydrogenase, and -ketoglutarate dehydrogenase. The
dehydrogenases are activated by Ca2+, which increases in
concentration during muscular contraction and secretion, when
there is increased energy demand.
contd ..
• --Increased [ATP]/[ADP] and [NADH]/[NAD+]
ratios inhibit PDH and the first three reactions.
• There is allosteric inhibition of citrate synthase by
ATP and long-chain fatty acyl-CoA.
The availability of oxaloacetate, as
controlled by malate dehydrogenase, depends
on the [NADH]/[NAD+] ratio.
The concentration of oxaloacetate
controls the rate of citrate formation.
Regulation of Iso-Citrate dehydrogenase:
In well fed state:
Inhibit Glycolysis
ATP
(-)
NADH
Enters fatty acid
synthesis
Accumulation of
Citrate
Iso-citrate
(-)
Isocitrate
Dehydrogenase
(-)
Alpha ketoglutarate
In well fed state, increase in ATP and NADH will inhibit isocitrate dehydrogenase leading
to accumulation of citrate. citrate will enter cytosol and inhibit Glycolysis and activates
fatty acid synthesis.
Regulation of citric acid cycle
• Citrate synthase
• Iso-citrate
dehydrogenase
• Alpha keto glutarate
dehydrogenase
• Aconitase – is inhibited by fluoroacetate (noncompetitive inhibition)
• -ketoglutarate dehydrogenase is inhibited
by Arsenite (non-competitive inhibition)
• Succinate dehydrogenase is inhibited by
Malonate (competitive inhibition)
• Beriberi , Wernicke’s encephalopathy and Korsakoff’s psychosis (WK
syndrome)in Thiamine deficiency is due to failure of TCA cycle (
Pyruvate dehydrogenase and - ketoglutarate dehydrogenase)
• Symptoms: confabulation, nystagmus (ophthalmoplegia), ataxia
• Congenital deficiency of Pyruvate dehydrogenase – Lactic acidosis
and neurodeficit.
• Congenital deficiency of Pyruvate carboxylase – OAA is deficient –
failure of sparking of TCA – severe mental retardation , lactic
acidosis, hypoglycemia
• TCA cycle enzyme deficiencies are extremely rare.
• During a myocardial infarction , the oxygen supply to
an area of the heart is dramatiocally reduced , forcing
the cardiac myocytes to switch to anaerobic
metabolism.Under these conditions , which of the
following enzymes would be activated by increasing
intracellular AMP?
A. Succinate dehydrogenase
B. PFK1
C. GLUCOKINASE
D. PDH
E. LDH
Which of the following is required for
cholesterol synthesis in hepatocytes?
•
•
•
•
•
A. Citrate shuttle
B. Glycerphosphate shuttle
C. Malate-Aspartate shuttle
D. Carnitine shuttle
E. Adenine nucleotide shuttle
• A 55 year old alcoholic was brought to the emergency department
by his friends. During their usual nightly gathering at the local bar,
he had passed out and they had been unable to revive him.
• The physician ordered an injection of thiamine followed by
overnight parental glucose. The next morning the patient was alert
and serum thiamine was normal and blood glucose was 73mg/dl.
• The IV line was removed and he was taken home. At the time of
discharge from hospital which of the following proteins would have
no significant physiological activity in this patient?
–
–
–
–
–
Malate dehydrogenase
Glucokinase
GLUT 1 transporter
PFK-1
Glucose 6 PO4 dehydrogenase

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