Transcript 213 lactate dehydrog..

Lactate Dehydrogenase
Prof. Dr. Laila Fadda
Formation of Acetyl CoA and lactate from Pyruvate
O2
Pyruvate dehydrogenase
complex
O2
Acetyl Co A
1. It is an enzyme which catalyzes the reaction
Lactate  Pyruvate
2. This reaction helps the re-oxidation of NADH, H+ into NAD+
3. It has 5 isoenzymes: LD1, LD2, LD3, LD4 and LD5.
4. Medical importance:
5. Estimation of the activity of lactate dehydrogenase enzyme
helps in the diagnosis of heart and liver diseases:
a) LD1: Elevated in some heart diseases e.g. myocardial
infraction.
b) LD5: Elevated in some liver diseases as acute viral hepatitis.
Regulation of glycolvsis:

The rate of glycolysis is regulated by controlling of
the 3 irreversible enzymes (key enzymes). These
enzymes catalyze what is called committed reactions
of the pathway. These enzymes are glucokinase
(hexokinase), phospho-fructokinase-1 and pyruvate
kinase.
1. Hormonal regulation:
a) Insulin: Stimulates synthesis of all key enzymes of glycolysis.
It is secreted after meal (in response to high blood glucose
level).
b) Glucagon: Inhibits the activity of all key enzymes of glycolysis.
It is secreted in response to low blood glucose level.
2. Energy regulation:
a) High level of ATP inhibits PFK-1 and pyruvate kinase.
b) High level of ADP and AMP stimulate PFK-1.
3. Substrate regulation:
a)
Glucose-6-phosphate inhibits hexokinase (and not glucokinase).
b) Citrate inhibits phosphofructokinase-1.
d) Fructose 1,6 bisphosphate stimulates pyruvate kinase.
Lactate:
Sources and fate of lactate:
a) Sources:

From glycolysis especially in RBCs due to
absence of mitochondria and muscle during
exercises due to oxygen lack.
2) Conversion into pyruvate: If oxygen gets available, lactate is
converted into pyruvate, which proceeds into Krebs' cycle.
3) Lactate may be accumulated in muscles causing muscle
fatigue.
4) Lactate may be excreted in urine and sweat.
Clinical aspects of glycolvsis:
1. There are many diseases associated with impaired glycolysis.
They include:
1. Pyruvate kinase deficiency.
2. Hexokinase deficiency.
3. Lactic acidosis.
2) Pyruvate kinase (PK) deficiency:
a) This leads to excessive hemolysis of RBCs
hemolytic anemia.
leading to
b) Genetic deficiency of PK enzyme causes decrease the rate of
glycolysis and decrease production of ATP.
c) ATP is required for Na' -K' ATPase, which is important for
stability of RBCs.
3. Hexokinase deficiency:
Leads to hemolytic anemia due to decrease ATP production.
The mechanism is similar to that of PK deficiency.
4. Lactic acidosis:
a) Definition and mechanism of lactic acidosis:
1) It is the lowered blood pH and bicarbonate levels due to increased
blood lactate above normal level.
OH
CH3-C -- COOH + NaHCO3
H2O
OH
CH3-C-COONa
+
H2CO3 CO2 +
Lactate+Sodium bicarbonate
Sodium lactate+Carbonic acid
2) This depletes bicarbonate ↓ pH of blood Lactic acidosis which
if severe may lead to coma.
b) Causes of lactic acidosis:
It results from increased formation or decreased utilization of
lactate.
1) Increased formation of lactate as in severe muscular exercises.
2) Decreased utilization of lactate in tissues: it occurs in cases of
anoxia or lack of oxygen. This is because oxygen is essential for
conversion of lactate into pyruvate, which proceeds into acetyl
CoA, and Krebs' cycle.
 Tissue
anoxia may occur in conditions that impair blood flow e.g.
myocardial infraction, angina pectoris, respiratory disorders, and
anemia.
3) Phenformin (cidophage): is oral hypoglycemic, causing excessive
anaerobic oxidation of glucose and excess lactate production.
II) Mitochondrial pathway for glucose oxidation:
A. Introduction: Complete oxidation of glucose occurs in both
cytoplasm (glycolysis) and mitochondria (Krebs' cycle) In the
presence of O2 pyruvate (the product of glycolysis) passes by
special pyruvate transporter into mitochondria which proceeds
as follows:
1. Oxidative decarboxylation of pyruvate into acetyl CoA.
2. Acetyl CoA is then oxidized completely to CO2, H2O through
Krebs' cycle.
B. Oxidative decarboxylation of pvruvate to acetvl
coenzvme A (= acetyl CoA):
1. Enzyme: Pyruvate dehydrogenase (PHD) complex.
a) This enzyme complex contains 4 subunits, which
catalyze the reaction in 4 steps.
b) This enzyme needs 5 coenzymes (all are vitamin B
complex derivatives):
1) Vitamin B1 = Thiamin diphosphate = TPP.
2) Lipoic acid = L -SH.-SH (reduced), L-s-s (oxidized).
3) Coenzyme A = CoASH.
4) Flavin adenine dinucleotide = FAD.
5) Nicotinamide adenine dinucleotide = NAD+
c) Location: PDH is located within the mitochondrial
matrix.
1) Energy production: (3ATP):

Oxidative decarboxylation of pyruvate to acetyl CoA
produces one molecule of NADH,H+. This produces
3
ATP
molecules
phosphorylation
through
respiratory
chain
2) Regulation of oxidative decarboxylation (PHD):
+ Insulin
Pyruvate
_ Calcium
Pyruvate DH complex
+NAD+
_Acetyl CoA
_ NADH+H
+CoASH
H2O
_ATP
a) Factors stimulating [+] PHD:
1) Pyruvate.
2) CoASH.
3) NAD+
4) Insulin hormone.
b) Factors inhibiting [-] PHD:
1) NADH,H+
2) ATP.
3) Acetyl CoA.
4) Calcium ions.

Kreb's cycle: [also known as citric acid cycle
(CAC) or tricarboxylic acid cycle (TCA) or
catabolism of acetyl CoA]:
1. Definition: TCA is a series of reactions in which
acetyl CoA is oxidized into CO2 +
energy.
2. Location: Mitochondria.
H2O and
The Citric Acid
(Krebs) Cycle
consists of eight
steps
Fig. 9.11
Tricarboxylic Acid Cycle
For oxidation of Acetyl CoA. •
Common pathway for final oxidation of proteins, carbohydrates and lipids. •
Occurs in mitochondrial matrix of all cells. •
Acetyl CoA + 3 NAD+ + FAD + GDP + Pi + 2H2O 
Reactions
 2CO2 + 3 NADH + FADH2 + GTP + 2H+ + CoA
3. Steps:
a) The enzymes of TCA cycle are present in the mitochondrial
matrix either free or attached to the inner surface of the
mitochondrial membrane.
b) The cycle is started by acetyl CoA (2 carbons) and oxaloacetate
(4 carbons) to form citrate (6 carbons). It ends by oxaloacetate
(4 carbons). The difference between the starting compound (6
carbons) and the ending compound (4 carbons) is 2 carbons
that are removed in the form of 2 CO2. These 2 carbons are
derived from acetyl CoA. For this reasons acetyl CoA is
completely catabolized in TCA and never gives glucose.
4. Energy production of TCA: (Energy catabolism of
acetyl CoA):
a) Oxidation of one molecule of acetyl CoA in TCA
produces 12 ATP molecules, 11 by respiratory chain
phosphorylation
and
1
phosphorylation as follows:
by
Substrate
level
Enzyme
Isocitrate degydrogenase
Method of ATP production
Oxidation
of
NADH+H
No. of ATP
by
3 ATP
respiratory chain phosph-orylation
α-Ketoglutarate
Oxidation
degydrogenase
respiratory chain phosphorylation
Succinyl CoA thiokinase
Substrate level phosphorylation
of
NADH+H
by
3 ATP
1 ATP
Succinate degydrogenase Oxidation of FADH by respiratory
2 ATP
chain phosphorylation
Malate degydrogenase
Oxidation
of
NADH+H
by
3 ATP
respiratory chain phosphorylation
Total =
12 ATP
b) Energy production of complete oxidation of one
molecule of glucose:
Glucose oxidation 36 or 38 ATP
Pyruvate oxidation 15ATP.
Acetyl CoA 12 ATP
5. Oxidative decarboxylation of α-ketoglutarate to succinyl
CoA
It is similar to the conversion of pyruvate to acetyl CoA.
a) Enzymes:α- ketoglutarate dehydrogenase complex.
b) Coenzymes: 5: TPP, Lipoic acid, CoASH, FAD and NAD+.
6. Functions (significance) of TCA:
The cycle is amphibolic i.e. it has catabolic (breakdown)
and anabolic (formation) functions.
Energy: 12 ATP
Catabolic functions: Oxidation of carbohydrate,
lipids and proteins
Anabolic function: Formation of:
 Amino
acids
 Glucose
 Heme
 Fatty
 CO2
acid and cholesterol
and over-controlling in his relation with
the patient.
a) Production of energy (12 ATP).
b) Catabolic functions: TCA is the final common
pathway for oxidation of carbohydrates, fats and
proteins (amino acids).
c) Anabolic functions: Formation of:
1) Amino acids:
-Ketoglutarate Transamination Glutamate.
Oxaloacetate
Transamination Aspratate.
2) Glucose: e.g.
Ketoglutarate Gluconeogenesis Glucose.
3) Heme synthesis:
Succinyl CoA
Heme.
4) Fatty acid and cholesterol:
Citrate (diffuse to cytoplasm) → Oxaloacetate + Acetyl
CoA → Fatty acid and cholesterol.
5) CO2 produced is used in the following (CO2 fixation)
reactions:

Pyruvate + CO2 GIVE Oxaloacetate by Gluconeogenesis
give Glucose.

Ammonia + ATP +CO2 give Carbamoyl phosphate give Urea
and pyrimidine

Formation of C6 of purine

Synthesis of H2CO3 / HCO, buffer.
The electron
transport chain
8. Regulation of citric acid cycle:

The key enzymes are citrate synthase dehydrogenase and ketoglutarate dehydrogenase:
a) Citrate synthase:
1) Stimulated by acetyl CoA, oxaloacetate, ADP and NAD+.
2 ) Inhibited by long chain acyl CoA, citrate, succinyl CoA, ATP
and NADH, H+
b) Isocitrate dehydrogenase and -ketoglutarate dehydrogenase:
1) Stimulated by NAD+, ADP.
2) Inhibited by NADH, H' and ATP
10. Energy production at substrate level in glucose oxidation:
a) The removal of hydrogen atoms from a compound is accompanied
by a release of energy. If this energy is captured in phosphate or
sulfate bonds, it will produce high-energy compounds.
b) The high energy compounds formed by glucose oxidation are:
1) Glyceraldhyde-3-p 1,3 BPG (phosphate bond).
2) Phosphoglycerate phosphoenol pyruvate (phosphate bond).
3) Pyruvate Acetyl CoA (sulfate bond(.
4) -Ketoglutarate Succinyl CoA (sulfate bond).