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Carbohydrate Metabolism — Viva Summary

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1. Overview

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2. Glycolysis

Two Phases:

• Glucose → Glucose-6-phosphate → Fructose-6-phosphate → Fructose-1,6-bisphosphate → 2× Glyceraldehyde-3-phosphate (G3P)

• G3P → 1,3-bisphosphoglycerate → 3-phosphoglycerate → 2-phosphoglycerate → Phosphoenolpyruvate (PEP) → Pyruvate

Key Regulated Enzymes (Irreversible Steps):

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3. Fate of Pyruvate

• Cofactors: Thiamine (B₁), Lipoic acid, FAD (B₂), NAD⁺ (B₃), CoA (B₅)
• Mnemonic: "The Lovely Foolish Nancy Comes"
• Inhibited by: NADH, Acetyl-CoA, ATP
• Activated by: NAD⁺, CoA, AMP, Ca²⁺

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4. TCA Cycle (Krebs / Citric Acid Cycle)

Steps and Enzymes:

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5. Oxidative Phosphorylation

• Each NADH → 2.5 ATP
• Each FADH₂ → 1.5 ATP

• Glycolysis: 2 ATP + 2 NADH (≈5 ATP via shuttle) = ~7
• PDC: 2 NADH = ~5 ATP
• TCA: 6 NADH (15) + 2 FADH₂ (3) + 2 GTP = 20 ATP
• Grand total: ~30–32 ATP

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6. Glycogenesis (Glycogen Synthesis)

1. Glucose → Glucose-6-P — Glucokinase/Hexokinase
2. Glucose-6-P → Glucose-1-P — Phosphoglucomutase
3. Glucose-1-P + UTP → UDP-Glucose + PPi — UDP-glucose pyrophosphorylase
4. UDP-Glucose added to glycogen chain (α-1,4 bonds) — Glycogen synthase (key enzyme)
5. Branching at every 8–10 residues (α-1,6 bonds) — Branching enzyme (amylo-4,6-transglucosidase)

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7. Glycogenolysis (Glycogen Breakdown)

1. Glycogen → Glucose-1-P — Glycogen phosphorylase (cleaves α-1,4 bonds) — key enzyme
2. Debrancher enzyme removes α-1,6 branches → free glucose + glucose-1-P
3. Glucose-1-P → Glucose-6-P — Phosphoglucomutase
4. Liver only: Glucose-6-P → Free glucose — Glucose-6-phosphatase (releases glucose into blood)
   • Muscle lacks glucose-6-phosphatase → glucose-6-P enters glycolysis directly

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8. Gluconeogenesis

• Pyruvate carboxylase requires biotin (CO₂ carrier) and is activated by acetyl-CoA
• PEPCK requires GTP

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9. Pentose Phosphate Pathway (PPP)

• Generates NADPH (for lipid synthesis, antioxidant defense via glutathione)
• Generates Ribose-5-phosphate (for nucleotide synthesis)
• Accounts for 5–10% of hepatic glucose metabolism

• Irreversible, rate-limiting
• Deficiency → hemolytic anemia on oxidant exposure (primaquine, fava beans)

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10. Hormonal Regulation at a Glance

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11. Quick-Reference: Key Enzymes Summary

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12. Clinically Important Points for Viva

• Lactate acidosis: From anaerobic glycolysis; e.g., shock, metformin overdose
• G6PD deficiency: X-linked; hemolysis with oxidant drugs (primaquine, dapsone)
• Von Gierke disease: Glucose-6-phosphatase deficiency → can't release glucose from liver → severe fasting hypoglycemia
• McArdle disease: Muscle glycogen phosphorylase deficiency → muscle cramps on exercise
• Pompe disease: Lysosomal acid maltase (α-1,4 glucosidase) deficiency → glycogen accumulation in all organs
• Pyruvate dehydrogenase deficiency: Lactic acidosis + neurological signs; treat with thiamine
• Fructose-2,6-bisphosphate: Master regulator — stimulates PFK-1 (glycolysis), inhibits FBPase-1 (gluconeogenesis); made by PFK-2, activated by insulin

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Enzymes — Viva Summary

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1. Definition & General Properties

• Enzymes are biological catalysts — mostly proteins (except ribozymes, which are RNA)
• They accelerate reaction rates by lowering the activation energy (ΔG‡) without being consumed
• They do not alter the equilibrium constant (Keq) or the standard free energy change (ΔG°) of a reaction
• They are highly specific — substrate specificity and reaction specificity
• They emerge unchanged at the end of the reaction

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2. Classification (IUB System — 6 Classes)

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3. Active Site & Substrate Binding

• Active site: Surface cleft or pocket on enzyme where substrate binds and catalysis occurs
• Contains: binding residues (hold substrate) + catalytic residues (break/make bonds)
• Active site provides: proximity, orientation, acid-base catalysis, electrostatic stabilization, covalent intermediates

Two Models of Enzyme-Substrate Binding:

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4. Enzyme Kinetics

Michaelis-Menten Equation:

Lineweaver-Burk Plot (Double Reciprocal):

• X-intercept = −1/Km
• Y-intercept = 1/Vmax
• Slope = Km/Vmax

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5. Factors Affecting Enzyme Activity

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6. Enzyme Inhibition

A. Reversible Inhibition

• Competitive: Statins (HMG-CoA reductase), Methotrexate (DHFR), Aspirin (COX), Sulfonamides (DHPS)
• Noncompetitive: Heavy metals (Pb, Hg) binding away from active site

B. Irreversible Inhibition

• Inhibitor forms covalent bond with enzyme → permanent inactivation
• Examples:
  • Aspirin → irreversibly acetylates COX-1/COX-2 (cyclooxygenase)
  • Organophosphates → irreversibly inhibit acetylcholinesterase (AChE)
  • Lead (Pb) → reacts with –SH groups of cysteine; inhibits ferrochelatase (blocks heme synthesis)
  • Penicillin → irreversibly inhibits transpeptidase (cell wall synthesis)
  • Sarin (nerve gas) → irreversibly inhibits AChE

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7. Regulation of Enzyme Activity

A. Allosteric Regulation

• Effector binds allosteric site (away from active site) → conformational change
• Positive effector (activator): ↑ enzyme activity
• Negative effector (inhibitor): ↓ enzyme activity
• Allosteric enzymes often show sigmoidal kinetics (not Michaelis-Menten) — described by the Hill equation
• Hill coefficient (n): n > 1 = positive cooperativity; n = 1 = no cooperativity
• Example: PFK-1 — activated by AMP/fructose-2,6-bisP; inhibited by ATP/citrate

B. Covalent Modification (Phosphorylation/Dephosphorylation)

• Kinases add phosphate (ATP → ADP) — phosphorylation
• Phosphatases remove phosphate — dephosphorylation

C. Zymogen Activation (Proteolytic Cleavage)

• Enzymes secreted as inactive precursors (zymogens) → activated by cleavage
• Examples:

D. Induction / Repression (Gene-Level)

• Hormones (insulin, glucagon, cortisol) regulate gene transcription of enzymes
• Example: Glucagon induces PEPCK (gluconeogenesis); Insulin induces glucokinase, fatty acid synthase

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8. Cofactors & Coenzymes

• Many enzymes require non-protein components for activity

Key Coenzymes (Vitamin Derivatives):

Key Metal Ion Cofactors:

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9. Isoenzymes (Isozymes)

• Multiple forms of the same enzyme catalyzing the same reaction but differing in:
  • Physical/chemical properties (electrophoretic mobility, Km, pH optimum)
  • Tissue distribution
  • Encoded by different genes or different subunit combinations

Clinically Important Isoenzymes:

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10. Enzymes as Diagnostic Markers

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11. Enzyme Deficiency Diseases (Viva High-Yield)

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12. Quick Summary — Key Regulatory Concepts

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Role of Cytochrome P450 (CYP450) in Drug Interactions

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1. What is Cytochrome P450?

R-H + O₂ + NADPH + H⁺ → R-OH + H₂O + NADP⁺

• Lippincott Illustrated Reviews: Biochemistry, 8th ed., p. 437
• Goodman & Gilman's Pharmacological Basis of Therapeutics, p. 120

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2. Location and Major CYP Isoforms

• Goodman & Gilman's Pharmacological Basis of Therapeutics, p. 121 (Table 5-2)
• Katzung's Basic and Clinical Pharmacology, 16th ed., p. 96

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3. Role in Drug Biotransformation (Phase I)

Phase I reactions catalyzed by CYP450:

• Katzung's Basic and Clinical Pharmacology, 16th ed., pp. 96-97
• Lippincott Illustrated Reviews: Biochemistry, 8th ed., p. 437

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4. Drug Interactions via CYP450

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A. CYP450 Inhibition

• Grapefruit juice inhibits CYP3A4 in intestinal wall → blood levels of statins (simvastatin), felodipine, and cyclosporine increase dramatically → risk of myopathy and toxicity.
• Ritonavir (HIV drug) is a potent CYP3A4 inhibitor — used intentionally to "boost" levels of other protease inhibitors (pharmacokinetic boosting).
• Katzung's Basic and Clinical Pharmacology, 16th ed., p. 97
• Goodman & Gilman's Pharmacological Basis of Therapeutics, p. 122

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B. CYP450 Induction

• Rifampicin (TB drug) induces CYP3A4 → rapidly metabolizes oral contraceptives → contraceptive failure and unintended pregnancy.
• Katzung's Basic and Clinical Pharmacology, 16th ed., p. 98
• Stahl's Essential Psychopharmacology, p. 55

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C. Prodrug Activation / Altered Activation

• Goodman & Gilman's Pharmacological Basis of Therapeutics, p. 124
• Katzung's Basic and Clinical Pharmacology, 16th ed., p. 97

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5. Genetic Polymorphism and Drug Interactions

• Stahl's Essential Psychopharmacology, p. 54
• Goodman & Gilman's Pharmacological Basis of Therapeutics, p. 121

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6. CYP450 and Paracetamol (Acetaminophen) Toxicity — Classic Example

• At therapeutic doses: CYP2E1 produces a small amount of the toxic metabolite NAPQI, which is safely neutralized by glutathione.
• In overdose or chronic alcoholism (CYP2E1 induced): Glutathione is depleted → NAPQI accumulates → centrilobular hepatic necrosis.
• Treatment: N-acetylcysteine (NAC) replenishes glutathione.
• Lippincott Illustrated Reviews: Biochemistry, 8th ed., p. 437
• Katzung's Basic and Clinical Pharmacology, 16th ed., p. 97

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Summary Table

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Sources with Page Numbers

• Purchase: Available on Amazon India, Flipkart, or medical bookstores — the 10th edition is widely stocked and reasonably priced (~₹600-900).
• Jaypee Brothers (the publisher) sells it directly at jaypeedigital.com — they also offer a digital/eBook version.
• College library — most medical college libraries stock Vasudevan; you can borrow or photocopy specific chapters for personal study.
• Google Play Books / Kindle — check if a legal digital edition is available on these platforms.
• Jaypee app — Jaypee Brothers has a mobile app with licensed access to their medical titles.

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• Lippincott Illustrated Reviews: Biochemistry, 8th ed. — covers liver function tests and enzyme markers
• Harper's Illustrated Biochemistry, 32nd ed. — covers isoenzymes and clinical significance
• Henry's Clinical Diagnosis and Management by Laboratory Methods — covers clinical enzyme diagnostics in detail

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Liver Enzymes and Their Importance in Clinical Diagnosis

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1. What are Liver Enzymes?

Important note: These are more accurately called markers of hepatocyte injury rather than true "liver function tests," since they reflect cell damage, not synthetic or metabolic function of the liver.

• Quick Compendium of Clinical Pathology, 5th ed., p. 2

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2. Major Liver Enzymes

A. Aminotransferases (Transaminases)

i. Alanine Aminotransferase (ALT) — formerly SGPT

• Location: Primarily cytoplasm of hepatocytes (liver-specific)
• Normal range: 7–56 U/L (males slightly higher than females)
• Reaction: Alanine + α-Ketoglutarate ⇌ Pyruvate + Glutamate (requires PLP/Vitamin B6)
• Significance: More specific for liver disease than AST; ALT elevation almost always indicates hepatic pathology
• ALT levels increase linearly with BMI; higher in males than females

ii. Aspartate Aminotransferase (AST) — formerly SGOT

• Location: Found in liver, cardiac muscle, skeletal muscle, kidney, brain, lung, pancreas (in decreasing order)
• ~80% of hepatic AST is concentrated in mitochondria; rest is cytoplasmic
• Normal range: 10–40 U/L
• Reaction: Aspartate + α-Ketoglutarate ⇌ Oxaloacetate + Glutamate (requires PLP)
• Significance: More sensitive (liver has large amounts of AST) but less specific than ALT
• Strenuous exercise and rhabdomyolysis can also raise AST
• Lippincott Illustrated Reviews: Biochemistry, 8th ed., pp. 703–705
• Quick Compendium of Clinical Pathology, 5th ed., pp. 2–3

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B. Alkaline Phosphatase (ALP)

• Location: Liver (bile canalicular membrane), bone, intestine, placenta, kidney
• Normal range: 44–147 U/L (higher in children due to bone growth)
• Significance: Raised in cholestatic liver disease (biliary obstruction, intrahepatic cholestasis) and bone disease
• ALP isoforms can be separated electrophoretically to distinguish hepatic from bone origin

• Quick Compendium of Clinical Pathology, 5th ed., p. 3

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C. Gamma-Glutamyl Transferase (GGT)

• Location: Liver, kidney, pancreas, intestine (liver is main serum source)
• Normal range: 9–48 U/L
• Key uses:
  • Most sensitive marker of hepatobiliary disease
  • Elevated in alcohol abuse (even without liver damage — GGT is induced by alcohol)
  • Useful to confirm that raised ALP is of hepatic (not bone) origin — if ALP ↑ and GGT ↑ = liver; if ALP ↑ and GGT normal = bone
  • Raised in cholestasis, fatty liver, hepatitis, drug-induced liver injury
• Lippincott Illustrated Reviews: Biochemistry, 8th ed., p. 705

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D. Lactate Dehydrogenase (LDH)

• Location: Widely distributed — liver, heart, RBCs, muscle, kidney, lung
• Isoforms:
  • LDH 1 & 2 — heart, RBCs, kidney
  • LDH 4 & 5 — liver and skeletal muscle
• Normal serum pattern: LDH2 > LDH1 > LDH3 > LDH4 > LDH5
• Raised LDH4 & LDH5 = liver damage or skeletal muscle injury
• LDH is non-specific; now largely replaced by more specific markers
• Still useful in: hemolysis, megaloblastic anemia, lymphoma, leukemia, germ cell tumors (primary marker for seminoma)
• Quick Compendium of Clinical Pathology, 5th ed., p. 2

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E. 5'-Nucleotidase (5'-NT)

• Specific to liver biliary canaliculi
• Raised in cholestatic liver disease
• More liver-specific than ALP; useful when ALP is raised to confirm hepatic origin
• Not affected by bone disease or pregnancy

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3. Clinical Importance — Pattern Recognition

Pattern 1 — Hepatocellular Damage (Hepatitis, Toxic Injury)

In severe viral hepatitis and toxic injury (e.g., Amanita phalloides poisoning), ALT can rise dramatically first, followed by rising bilirubin as hepatocellular damage progresses.

• Lippincott Illustrated Reviews: Biochemistry, 8th ed., p. 704

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Pattern 2 — Cholestatic Disease (Biliary Obstruction)

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Pattern 3 — Alcoholic Liver Disease

The De Ritis ratio (AST:ALT) is a key diagnostic tool:

• ALT > AST = most viral/toxic hepatitis
• AST > ALT (ratio >2:1) = alcoholic liver disease
• AST > ALT (ratio >3:1) = strongly suggests alcoholic hepatitis
• Wilson's disease and cirrhosis can also show reversal of the ratio

• Quick Compendium of Clinical Pathology, 5th ed., p. 3
• Lippincott Illustrated Reviews: Biochemistry, 8th ed., p. 705

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Pattern 4 — Differentiating Liver from Bone (Raised ALP)

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4. Summary Table — Liver Enzymes at a Glance

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Sources & Page Numbers

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Therapeutic and Diagnostic Uses of Enzymes

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PART A — DIAGNOSTIC USES OF ENZYMES

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1. Enzymes in Cardiac Disease

• CK isoforms: CK-MM (skeletal muscle), CK-MB (heart), CK-BB (brain)
• CK-MB > 6% of total CK = diagnostic of MI
• LDH "flipped ratio" (LDH1 > LDH2) = hallmark of MI (normally LDH2 > LDH1)
• Strenuous exercise → ↑ CK-MM (not CK-MB) — differentiates from MI

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2. Enzymes in Liver Disease

• ALT > AST = viral hepatitis, toxic injury
• AST:ALT > 2:1 = alcoholic liver disease
• AST:ALT > 3:1 = strongly suggests alcoholic hepatitis

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3. Enzymes in Pancreatic Disease

• Serum lipase is the preferred test for acute pancreatitis today (more specific than amylase)
• Salivary amylase (S-type) elevated in parotitis (mumps) — not pancreatitis
• Pancreatic amylase (P-type) separated by electrophoresis

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4. Enzymes in Bone Disease

• ALP raised in bone disease but GGT is normal — distinguishes from liver-origin ALP
• Tartrate-resistant acid phosphatase (TRAP) is a marker of hairy cell leukaemia

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5. Enzymes in Muscle Disease

• CK is the most sensitive marker for muscular dystrophy — elevated even before symptoms appear
• In rhabdomyolysis: CK-MM, AST, ALT, and myoglobin all rise

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6. Enzymes in Prostate Disease

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7. Enzymes as Diagnostic Tools in the Lab (ELISA)

• Horseradish peroxidase and alkaline phosphatase are used as enzyme labels in ELISA (Enzyme-Linked Immunosorbent Assay) to detect antibodies, antigens, hormones, and drugs
• The enzyme-linked antibody produces a colour change proportional to the analyte concentration
• Used for: HIV diagnosis, hepatitis B surface antigen, pregnancy tests (hCG), drug levels, tumour markers

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Summary — Diagnostic Uses

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PART B — THERAPEUTIC USES OF ENZYMES

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1. Enzyme Replacement Therapy (ERT)

• β-Glucocerebrosidase is modified to expose mannose residues → targets macrophage mannose receptor → enzyme enters lysosomes → degrades accumulated glucocerebroside
• Results: normalization of anemia, platelet count, liver/spleen size, bone density
• Only ~1–5% of normal enzyme activity is needed to correct biochemical abnormalities
• Limitation: Does not cross blood-brain barrier → cannot treat neurological (type 2/3) Gaucher disease

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2. Thrombolytic Enzymes (Fibrinolytic Therapy)

• These are serine proteases that act on the fibrinolytic pathway
• tPA is preferred in stroke as it is clot-specific (less systemic bleeding risk than streptokinase)

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3. Digestive Enzyme Preparations

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4. Enzyme Inhibition as Therapy (Enzyme Inhibitors Used as Drugs)

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5. Enzymes in Wound Care and Debridement

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6. Enzymes in Cancer Treatment

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7. Enzymes in Other Clinical Uses

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Complete Summary Table

Diagnostic

Therapeutic

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Sources & Page Numbers