Biochemistry

Part III · Common Biology · Chapter Fourteen

Expect 8–12 questions: net ATP yield from glucose, enzyme inhibition types (competitive vs non-competitive), glycolysis regulatory enzymes (PFK-1 is the key control), Mitchell’s chemiosmotic theory, protein structure levels, essential amino acids (PVT TIM HALL), DNA vs RNA, reducing sugars, vitamin deficiency diseases, and biochemical tests (Biuret, Benedict’s, Molisch). Year-person-discovery items — Krebs, Mitchell, Sumner, Wöhler — are reliably tested.

Read · 80 min
Revise · 20 min
MCQs · 28

Syllabus Coverage

Biomolecules — carbohydrates, lipids, proteins, nucleic acids • Enzymes: structure, kinetics, inhibition, regulation • Metabolism: glycolysis, TCA cycle, electron transport chain & oxidative phosphorylation, pentose phosphate pathway, β-oxidation • Vitamins and cofactors • Nitrogen metabolism and urea cycle • Biochemical tests and detection methods.

14.1 Carbohydrates

Carbohydrates are the most abundant organic compounds on Earth. Their empirical formula is C_n(H₂O)_n, which led early chemists to regard them as “hydrates of carbon.” They serve as fuels, structural elements (cellulose, chitin), and informational scaffolds (nucleic-acid sugars).

Wöhler 1828 — urea from NH₄OCN, ending the vitalism doctrine · Berzelius 1838 — coined “proteins” · Emil Fischer 1891 — stereochemistry of sugars, D/L convention, Fischer projection · Haworth 1929 (Nobel 1937) — cyclic (pyranose/furanose) sugar structures

14.1.1 Monosaccharides

Monosaccharides are the simplest carbohydrates. Hexoses (C₆H₁₂O₆) include glucose, fructose, and galactose. Pentoses (C₅H₁₀O₅) include ribose (in RNA) and 2-deoxyribose (in DNA). Glucose and galactose are aldoses (aldehyde at C-1); fructose is a ketose (keto at C-2). In solution glucose cyclises through the hydroxyl at C-5 to form glucopyranose: the α-anomer has the C-1 –OH axial (down in Haworth), and the β-anomer has it equatorial (up). This anomeric difference determines whether the polymer is starch (α-linkages) or cellulose (β-linkages). D-sugars rotate the plane of polarised light rightward (dextrorotatory) in most cases; all natural sugars are of D-configuration by Fischer convention (hydroxyl on the right at the penultimate carbon).

14.1.2 Disaccharides

Two monosaccharides joined by a glycosidic bond (condensation, −H₂O). Key disaccharides:

Sucrose (table sugar) — glucose (C-1) + fructose (C-2) via α,β-1,2 linkage; non-reducing (both anomeric carbons blocked); hydrolysis by sucrase/invertase yields “invert sugar.”
Lactose (milk sugar) — galactose + glucose via β-1,4 linkage; reducing (free anomeric C of glucose); digested by lactase (deficiency = lactose intolerance).
Maltose (malt sugar) — glucose + glucose via α-1,4 linkage; reducing; product of starch digestion by amylase.
Cellobiose — glucose + glucose via β-1,4 linkage; reducing; structural disaccharide unit of cellulose.
Trehalose — glucose + glucose via α,α-1,1; non-reducing; major circulating sugar of insects; also in yeast and desiccation-tolerant plants.

Easy to confuse — Reducing vs Non-reducing Sugars

Reducing Sugar

Has a free anomeric (aldehyde/keto) carbon available to donate electrons to Cu²⁺. Detected by Fehling’s (brick-red Cu₂O) and Benedict’s (red/orange/yellow precipitate). Examples: glucose, fructose, galactose, lactose, maltose.

Non-reducing Sugar

Both anomeric carbons locked in the glycosidic bond; cannot reduce Cu²⁺ directly. Examples: sucrose, trehalose. Must be hydrolysed first. Negative Fehling’s; positive after acid hydrolysis.

14.1.3 Polysaccharides

Long chains of monosaccharide units joined by glycosidic bonds. Polysaccharides are the most important storage and structural macromolecules in biology.

**Table 14.1**Major polysaccharides — structure and function
Polysaccharide	Monomer	Linkage	Branching	Function / Location
Amylose	Glucose	α-1,4	None (linear)	Starch component; storage in plants
Amylopectin	Glucose	α-1,4 + α-1,6	Every 24–30 residues	Starch component; 80% of starch
Glycogen	Glucose	α-1,4 + α-1,6	Every 8–12 residues	Animal + yeast storage; liver & muscle
Cellulose	Glucose	β-1,4	None (linear fibres)	Plant cell-wall structural element
Chitin	N-acetylglucosamine	β-1,4	None	Fungal cell wall; arthropod exoskeleton
Inulin	Fructose	β-2,1	None	Reserve in chicory, garlic, dahlia tubers
Agar	Agarose + agaropectin	β-1,3 / α-1,4	Sulphated	Red algae cell wall; microbiology gel medium

Exam Tip: Glycogen is more branched than amylopectin (branch every 8–12 residues vs 24–30). Iodine stain gives blue-black with amylose (helix traps I₃−), purple-brown with amylopectin/glycogen, and no colour with cellulose. The Molisch test (H₂SO₄ + α-naphthol) is positive for all carbohydrates — purple ring at the interface.

14.2 Lipids

Lipids are chemically heterogeneous biomolecules united by hydrophobicity (insoluble in water, soluble in organic solvents). They include fats, oils, phospholipids, waxes, steroids, and fat-soluble vitamins. The term was coined by Bloor (1943).

14.2.1 Fatty Acids

Fatty acids are long-chain carboxylic acids (generally C₁₂–C₂₂). Saturated fatty acids contain no double bonds: palmitic acid (C_16:0, hexadecanoic) and stearic acid (C_18:0, octadecanoic) are the most common. Unsaturated fatty acids contain one or more C=C double bonds. Oleic acid (C_18:1, Δ⁹ cis) is the most abundant monounsaturated fatty acid in nature (olive oil). Linoleic acid (C_18:2, ω-6) and α-linolenic acid (C_18:3, ω-3) are essential fatty acids — humans cannot synthesise them (lack Δ¹² and Δ¹⁵ desaturases). Arachidonic acid (C_20:4, ω-6) is conditionally essential and is the precursor to prostaglandins, thromboxanes, and leukotrienes.

Easy to confuse — Saturated vs Unsaturated vs Trans Fatty Acids

Saturated FAs

No C=C; straight chain; tightly packed; solid at room temperature (butter, ghee, coconut oil). Associated with increased LDL cholesterol. Palmitic, stearic.

Unsaturated FAs

One or more cis C=C; kinked chain; loosely packed; liquid at room temperature (vegetable oils). Oleic, linoleic, linolenic. Trans fats: unsaturated but trans-configuration (partial hydrogenation); behave like saturated; raise LDL, lower HDL; harmful to cardiovascular health.

14.2.2 Triglycerides, Phospholipids, and Steroids

Triglycerides (triacylglycerols) = glycerol esterified with three fatty acids. They are the main energy storage lipid in adipose tissue. The iodine number measures degree of unsaturation (grams of I₂ absorbed per 100 g fat); the saponification number measures average chain length (moles of KOH per gram of fat; smaller FA = larger saponification number). Phospholipids replace one FA with a phosphate head group; they are amphipathic and form the bilayer of all cell membranes (fluid mosaic model, Singer & Nicolson 1972). Steroids share the four-ring (cyclopentanoperhydrophenanthrene) sterane skeleton. Cholesterol is the precursor to all steroid hormones (cortisol, testosterone, oestradiol), bile acids, and vitamin D. Waxes are esters of long-chain FA + long-chain alcohol; present on leaf cuticle and arthropod exoskeletons (beeswax = myricyl palmitate).

HP angle: Sea buckthorn (Hippophae rhamnoides), found wild in Lahaul-Spiti and Kinnaur, is one of the richest plant sources of omega-3 and omega-7 (vaccenic) fatty acids and vitamin C (>400 mg/100 g). Trout fish (Oncorhynchus mykiss), farmed in HP rivers, is rich in EPA and DHA (ω-3). Apple polyphenols (quercetin, chlorogenic acid) from HP orchards are potent antioxidants with current nutraceutical research.

14.3 Proteins & Amino Acids

Proteins are polymers of L-α-amino acids linked by peptide bonds (−CO–NH–). They are the most functionally diverse macromolecules: enzymes, structural scaffolds (collagen, keratin), transporters (haemoglobin), hormones (insulin), receptors, antibodies, and motors (myosin). The term “protein” was introduced by Berzelius (1838); the tetrahedral carbon chemistry was explained by van’t Hoff and Le Bel (1874).

14.3.1 Amino Acid Structure

Every standard amino acid has a central α-carbon bearing: (1) an amino group (–NH₂), (2) a carboxyl group (–COOH), (3) a hydrogen, and (4) a variable R group (side chain) that determines identity and properties. At physiological pH 7.4, the –NH₂ is protonated (–NH₃⁺) and –COOH is deprotonated (–COO⁻), making the molecule a zwitterion. Only glycine lacks chirality (R = H); all other amino acids are L-configuration by convention.

Fig. 14.1General structure of an α-amino acid (left) and the four broad R-group classifications (right). At physiological pH the molecule exists as a zwitterion: –NH₃⁺ and –COO⁻.

14.3.2 Essential Amino Acids

Essential amino acids cannot be synthesised by the human body (or are synthesised in quantities insufficient for normal growth) and must be obtained from diet. There are 8 essential in adults (with His and Arg essential in children / during growth):

Mnemonic — Essential Amino Acids (10 including His & Arg)

PVT TIM HALL

P = Phenylalanine V = Valine T = Threonine T = Tryptophan I = Isoleucine M = Methionine H = Histidine A = Arginine L = Leucine L = Lysine

14.3.3 Protein Structure Levels

Easy to confuse — Four Levels of Protein Structure

Primary

Linear sequence of amino acids in the polypeptide chain (determined by gene). Bond: peptide bond (covalent). Sanger 1953 sequenced insulin (first protein).

Tertiary

Overall 3-D fold of the single chain. Stabilised by hydrophobic interactions (major driving force), ionic bonds, H-bonds, and disulphide bridges (Cys–Cys, covalent). Anfinsen 1972 (Nobel) showed folding is encoded in primary sequence.

Secondary

Local regular structures: α-helix (3.6 residues per turn; H-bonds between C=O of residue n and N–H of n+4; right-handed helix; Pauling & Corey 1951) and β-pleated sheet (H-bonds between adjacent extended strands; parallel or antiparallel). Both stabilised by backbone H-bonds only.

Quaternary

Assembly of multiple polypeptide subunits. Example: haemoglobin = 2α + 2β subunits. Insulin = A + B chains. DNA polymerase. Collagen = triple helix. Stabilised by non-covalent interactions between subunits.

Denaturation disrupts secondary, tertiary, and quaternary structure (primary sequence intact) by heat, extreme pH, detergents, or heavy metals. Denaturation is often irreversible (egg white coagulation). Renaturation is possible for some small proteins (Anfinsen’s ribonuclease experiment). Chaperones (Hsp70, GroEL/GroES) assist folding in vivo.

Peptide Bond

A covalent amide bond formed between the α-carboxyl group of one amino acid and the α-amino group of another with release of water (condensation reaction). The C–N bond has partial double-bond character due to electron delocalisation, making the peptide bond planar and normally in the trans configuration. A chain of amino acids linked by peptide bonds is a polypeptide.

14.3.4 Biochemical Tests for Proteins and Amino Acids

**Table 14.2**Biochemical colour tests
Test	Reagent	Positive result	Detects
Biuret	NaOH + dilute CuSO₄	Purple/violet	Peptide bonds (proteins with ≥2 peptide bonds)
Ninhydrin	Ninhydrin solution, heat	Purple (Ruhemann’s purple)	Free α-amino acids; Pro gives yellow
Molisch	α-naphthol + conc. H₂SO₄	Purple ring at interface	All carbohydrates (universal CHO test)
Benedict’s	Sodium citrate + CuSO₄, heat	Brick-red/orange/yellow ppt.	Reducing sugars
Fehling’s	Fehling A + B, heat	Brick-red Cu₂O precipitate	Reducing sugars
Iodine	I₂/KI solution	Blue-black	Starch (amylose helix)
Sudan III/IV	Sudan dye	Red/orange staining	Lipids (fats and oils)
Xanthoproteic	Conc. HNO₃ + NaOH	Yellow → orange	Aromatic amino acids (Phe, Tyr, Trp)

14.4 Nucleic Acids — DNA, RNA, and Nucleotides

Nucleic acids are polymers of nucleotides linked by 3′→5′ phosphodiester bonds. They carry genetic information (DNA) and mediate its expression (RNA). The term “nuclein” was coined by Miescher (1869) who isolated it from pus-cell nuclei; later renamed nucleic acid by Altmann (1889). The double-helical structure of DNA was proposed by Watson and Crick (1953) using X-ray diffraction data from Franklin and Wilkins (Nobel to Watson, Crick, Wilkins 1962).

14.4.1 Nucleotide Structure

A nucleotide consists of three components: (1) a nitrogenous base, (2) a pentose sugar (ribose in RNA; 2′-deoxyribose in DNA), and (3) one or more phosphate groups. The nitrogenous bases fall into two families:

Purines (double-ring): Adenine (A) and Guanine (G) — shared by DNA and RNA.
Pyrimidines (single-ring): Cytosine (C) in both; Thymine (T) in DNA only; Uracil (U) in RNA only.

Mnemonic: Pur-ines = Pure Silver (PurinAG); pyrimidines cut (CUT — Cytosine, Uracil, Thymine). A nucleoside = base + sugar (no phosphate). ATP (adenosine triphosphate) is the universal energy currency; hydrolysis of the terminal phosphate anhydride bond releases ~7.3 kcal/mol (−30.5 kJ/mol).

Easy to confuse — DNA vs RNA Structure

DNA

Sugar: 2′-deoxyribose. Bases: A, G, C, T. Usually double-stranded antiparallel helix (B-form most common). Base pairing: A=T (2 H-bonds), G≡C (3 H-bonds). Chargaff’s rules: [A]=[T]; [G]=[C]; A+G = C+T (purine = pyrimidine). Stable; 2′-OH absent → resistant to alkaline hydrolysis. Localised in nucleus (+ mitochondria, plastids).

RNA

Sugar: ribose (2′-OH). Bases: A, G, C, U. Usually single-stranded (can form hairpin loops, stems). Less stable than DNA; 2′-OH makes it susceptible to alkaline hydrolysis. Types: mRNA (messenger), tRNA (transfer; cloverleaf; carries amino acid), rRNA (ribosomal; catalytic), miRNA/siRNA (regulatory), lncRNA (long non-coding).

**Table 14.3**DNA vs RNA — key comparison
Feature	DNA	RNA
Sugar	2′-Deoxyribose	Ribose
Unique base	Thymine (T)	Uracil (U)
Strandedness	Usually dsDNA	Usually ssRNA
Stability	More stable (no 2′-OH)	Less stable (2′-OH)
Location	Nucleus, mitochondria, chloroplasts	Nucleus, cytoplasm, ribosomes
Function	Genetic information storage	Gene expression (mRNA, tRNA, rRNA)
Transcription product?	Template	Product
Chargaff’s rule applies?	Yes: A=T, G=C	Not applicable (ss)

Exam Tip: Ribozymes are catalytic RNA molecules (e.g., self-splicing introns, peptidyl transferase activity of rRNA). This blurs the protein-only enzyme concept — expect one question on “which of the following is both catalytic and informational?” Answer: RNA (ribozyme). Thomas Cech and Sidney Altman shared Nobel 1989 for ribozyme discovery.

Miescher 1869 — isolated “nuclein” from white blood cells · Chargaff 1950 — A=T, G=C base-pairing rules · Watson & Crick 1953 — double-helix model (Nobel 1962) · Cech & Altman 1989 — RNA catalysis / ribozymes (Nobel) · Fire & Mello 2006 — RNA interference (Nobel)

14.5 Enzymes & Enzyme Kinetics

Enzymes are biological catalysts that accelerate chemical reactions by lowering activation energy (E_a) without being consumed. Almost all enzymes are proteins (with some notable RNA exceptions — ribozymes). The systematic study of enzymes began with Eduard Buchner (1897) who showed cell-free yeast extract could ferment sugar — proving catalysis is a molecular, not cellular, phenomenon (Nobel 1907). James Sumner (1926) crystallised urease from jack bean and demonstrated it was a protein (Nobel 1946).

Enzyme

A biological catalyst (nearly always a protein; sometimes RNA) that lowers the activation energy of a specific chemical reaction, thereby increasing its rate by factors of 10⁶–10¹² over the uncatalysed rate. Enzymes are not consumed; they emerge unchanged from each catalytic cycle. They provide a micro-environment on the active site that stabilises the transition state.

14.5.1 Active Site and Substrate Binding

The active site is a specific three-dimensional cleft or crevice in the enzyme, formed by amino acid residues that may be far apart in the primary sequence. Two models explain substrate binding:

Fig. 14.2Lock-and-key model (Fischer, 1894): active site is rigid and pre-complementary to the substrate shape. Induced-fit model (Koshland, 1958): the enzyme undergoes a conformational change on substrate binding, wrapping around it. Most modern evidence supports induced fit (e.g., hexokinase closes around glucose).

14.5.2 Michaelis-Menten Kinetics

The Michaelis-Menten equation (Michaelis & Menten 1913) describes enzyme kinetics for a simple one-substrate reaction: E + S ⇌ ES → E + P.

V_max = maximum reaction velocity when enzyme is saturated with substrate. K_m (Michaelis constant) = substrate concentration at half-maximum velocity. A low K_m means high affinity. The Lineweaver-Burk double-reciprocal plot (1/v vs 1/[S]) gives a straight line with slope = K_m/V_max, x-intercept = −1/K_m, and y-intercept = 1/V_max — very useful for distinguishing inhibition types.

14.5.3 Enzyme Inhibition

Easy to confuse — Competitive vs Non-competitive Inhibition

Competitive Inhibition

Inhibitor structurally resembles substrate and binds the active site. Can be overcome by increasing [S]. Effect: K_m increases (apparent, less affinity); V_max unchanged. Lineweaver-Burk: lines intersect at y-axis. Example: malonate inhibiting succinate dehydrogenase; statins inhibiting HMG-CoA reductase.

Non-competitive Inhibition

Inhibitor binds an allosteric (non-active) site, distorting enzyme shape. Cannot be overcome by increasing [S]. Effect: V_max decreases; K_m unchanged. Lineweaver-Burk: lines intersect at x-axis. Example: heavy metals (Pb²⁺, Hg²⁺) inhibiting enzymes with –SH groups; cyanide inhibiting cytochrome c oxidase.

Uncompetitive inhibition: inhibitor binds only the ES complex; both K_m and V_max decrease. Lineweaver-Burk: parallel lines (slope unchanged). Irreversible inhibition: covalent modification of active site. Examples: organophosphates (sarin, nerve agents) covalently inhibit acetylcholinesterase; aspirin (acetylsalicylate) irreversibly acetylates cyclooxygenase (COX-1/COX-2); penicillin acylates the transpeptidase (DD-transpeptidase) that cross-links peptidoglycan.

Easy to confuse — Allosteric Regulation vs Feedback Inhibition

Allosteric Regulation

Regulator molecule (activator or inhibitor) binds a separate allosteric site, causing a conformational change that alters catalytic activity. Allosteric enzymes often show sigmoidal (cooperative) kinetics rather than hyperbolic Michaelis-Menten. Key example: phosphofructokinase-1 (PFK-1) — activated by AMP/ADP, inhibited by ATP/citrate.

Feedback (End-product) Inhibition

The final product of a metabolic pathway inhibits an early enzyme in that pathway (usually allosterically). A specific form of allosteric inhibition. Classic example: threonine deaminase inhibited by isoleucine (end product of the Thr → Ile pathway). Also: CTP inhibiting aspartate transcarbamoylase (ATCase) in pyrimidine synthesis. Prevents overproduction of end products.

14.5.4 Cofactors, Coenzymes, and Prosthetic Groups

Many enzymes require non-protein accessory molecules for activity:

Cofactors — inorganic ions: Mg²⁺ (DNA polymerase, kinases), Zn²⁺ (carbonic anhydrase, alcohol dehydrogenase), Fe^2+/3+ (cytochromes, haemoglobin), Cu²⁺ (cytochrome c oxidase), Mn²⁺ (arginase), K⁺ (pyruvate kinase).
Coenzymes — organic, loosely bound, vitamin-derived: NAD⁺/NADH (from niacin/B₃), FAD/FADH₂ (from riboflavin/B₂), CoA (from pantothenic acid/B₅), TPP/thiamine pyrophosphate (from thiamine/B₁), pyridoxal phosphate (from B₆), biotin (B₇, CO₂ carrier in carboxylases), tetrahydrofolate (from folate/B₉), methylcobalamin (from B₁₂).
Prosthetic groups — tightly (covalently) bound: haem in cytochromes and haemoglobin, FAD in succinate dehydrogenase, biotin in pyruvate carboxylase.

A holoenzyme = apoenzyme (protein) + cofactor/coenzyme (active). An apoenzyme alone is catalytically inactive.

14.5.5 IUBMB Enzyme Classification

**Table 14.4**IUBMB enzyme classes (EC numbers)
Class	Name	Reaction type	Examples
EC 1	Oxidoreductases	Oxidation-reduction (transfer of H, O, or electrons)	Dehydrogenases, oxidases, reductases, peroxidases
EC 2	Transferases	Transfer of functional groups (acyl, methyl, glycosyl, phosphoryl)	Kinases, transaminases, polymerases
EC 3	Hydrolases	Hydrolysis (water as co-substrate)	Lipases, proteases, amylases, nucleases, phosphatases
EC 4	Lyases	Addition/removal of groups to/from double bonds (non-hydrolytic, non-oxidative)	Decarboxylases, aldolases, hydratases
EC 5	Isomerases	Intramolecular rearrangements (isomerisation)	Racemases, mutases, epimerases, isomerases
EC 6	Ligases	Bond formation coupled to ATP hydrolysis	Synthetases, carboxylases, aminoacyl-tRNA synthetases
EC 7	Translocases	Translocation of ions/molecules across membranes	ATP-synthase, Na⁺/K⁺-ATPase (added 2018)

Worked Example — Classify an Enzyme

“Hexokinase catalyses the transfer of a phosphate group from ATP to glucose, forming glucose-6-phosphate and ADP. To which IUBMB class does hexokinase belong?”

Answer: EC 2 — Transferase. It transfers a phosphoryl group (functional group) from ATP to glucose. Its sub-class is phosphotransferase (EC 2.7.1.1, hexokinase). Note: kinase = transferase, always.

14.6 Glycolysis & Fermentation

Glycolysis (from Greek: glykys = sweet; lysis = splitting) is the universal 10-step pathway that converts one molecule of glucose (C₆) to two molecules of pyruvate (C₃), occurring in the cytoplasm of all living cells. It is the oldest energy pathway, evolutionarily predating the mitochondrion. The pathway was established by Embden, Meyerhof, and Parnas in the 1930s and is often called the Embden–Meyerhof–Parnas (EMP) pathway.

Buchner 1897 — cell-free fermentation (Nobel 1907) · Embden, Meyerhof, Parnas 1930s — glycolysis pathway · Meyerhof Nobel 1922 (lactic acid / muscle work) · Warburg Nobel 1931 (respiratory enzymes; also Warburg effect in cancer cells)

Fig. 14.3The 10-step glycolytic (EMP) pathway. Steps 1–5 are the energy investment phase (2 ATP consumed); steps 6–10 are the energy payoff phase (4 ATP + 2 NADH produced per glucose, or 2× per triose). Net yield: 2 ATP + 2 NADH + 2 pyruvate. Starred enzymes (hexokinase, PFK-1, pyruvate kinase) are the major regulatory points; PFK-1 is the primary control valve.

Easy to confuse — Aerobic vs Anaerobic Respiration

Aerobic Respiration

Requires O₂ as final electron acceptor. Pyruvate → acetyl-CoA → Krebs cycle → ETC → 30–32 ATP per glucose. Complete oxidation of glucose to CO₂ + H₂O. Occurs in mitochondria (after glycolysis in cytoplasm).

Anaerobic Respiration / Fermentation

No O₂ needed. Pyruvate is converted to regenerate NAD⁺ (essential to keep glycolysis running). Two main routes: Lactate fermentation (animals, Lactobacillus): pyruvate + NADH → lactate + NAD⁺ [lactate dehydrogenase]. Ethanol fermentation (yeast Saccharomyces, plant roots in waterlogging): pyruvate → acetaldehyde (CO₂ released) → ethanol. Net: 2 ATP per glucose only. HP relevance: chhang and lugri (barley-based fermented beverages of Kinnaur/Lahaul) and chyangra (Himalayan goat milk fermented products) rely on lactic acid fermentation.

Easy to confuse — Glycolysis vs Gluconeogenesis

Glycolysis

Glucose → pyruvate. Net: 2 ATP produced. Occurs in all cells. Active when glucose/energy available. Regulatory enzymes: hexokinase, PFK-1, pyruvate kinase (activated by AMP/ADP; inhibited by ATP/citrate). Main site: cytoplasm.

Gluconeogenesis

Pyruvate + lactate + amino acids → glucose. Net: 4 ATP + 2 GTP consumed. Active in liver/kidney during fasting. Bypasses 3 irreversible glycolytic steps using unique enzymes: pyruvate carboxylase, PEPCK, fructose-1,6-bisphosphatase, glucose-6-phosphatase. Predominantly in liver cytoplasm + mitochondria. Cannot occur in muscle (no G6Pase).

14.7 Krebs (TCA) Cycle & Pyruvate Oxidation

Before entering the Krebs cycle, pyruvate from glycolysis is transported into the mitochondrial matrix and oxidatively decarboxylated by the pyruvate dehydrogenase complex (PDC) — a huge multi-enzyme assembly (E1/pyruvate decarboxylase, E2/dihydrolipoyl transacetylase, E3/dihydrolipoyl dehydrogenase) requiring TPP, lipoate, CoA, NAD⁺, and FAD as cofactors.

Pyruvate oxidation: Pyruvate + NAD⁺ + CoA → Acetyl-CoA + NADH + CO₂. Per pyruvate: 1 NADH, 1 CO₂, 1 Acetyl-CoA. Per glucose: 2 NADH, 2 CO₂ (2 pyruvates).

The citric acid (TCA / Krebs) cycle was elucidated by Hans Krebs in 1937 (Nobel Prize 1953). It operates in the mitochondrial matrix and is the central hub of aerobic catabolism, accepting acetyl-CoA from all major fuel sources.

Krebs 1937 — proposed citric acid (TCA) cycle (Nobel 1953) · Krebs & Henseleit 1932 — urea cycle · Lipmann 1945 — acetyl-CoA and CoA (Nobel 1953, shared with Krebs) · Ochoa — isolated several Krebs cycle enzymes (Nobel 1959)

Fig. 14.4The Krebs (TCA/citric acid) cycle. One turn (per acetyl-CoA) yields: 3 NADH, 1 FADH₂, 1 GTP, and 2 CO₂. Per glucose (2 turns): 6 NADH, 2 FADH₂, 2 GTP, 4 CO₂. OAA is regenerated at the end, making it catalytic. Citrate synthase and isocitrate dehydrogenase are the main regulatory enzymes.

Easy to confuse — Substrate-level vs Oxidative Phosphorylation

Substrate-level Phosphorylation

ATP synthesised directly from a high-energy substrate intermediate by enzyme catalysis. No membrane, no H⁺ gradient needed. Occurs in glycolysis (phosphoglycerate kinase, pyruvate kinase) and Krebs cycle (succinyl-CoA synthetase). Yields 2+2 = 4 ATP per glucose substrate-level.

Oxidative Phosphorylation

ATP synthesised by ATP synthase (Complex V) using the proton-motive force (proton gradient) across the inner mitochondrial membrane generated by the electron transport chain. Requires O₂ as final electron acceptor. Chemiosmotic theory (Mitchell 1961, Nobel 1978). Yields ~26–28 ATP per glucose (the bulk of aerobic ATP).

14.8 Electron Transport Chain & Oxidative Phosphorylation

The ETC is embedded in the inner mitochondrial membrane (IMM). NADH and FADH₂ donate electrons, which flow down an energy gradient through four major protein complexes and two mobile carriers, ultimately reducing O₂ to H₂O. The energy released by electron flow drives proton pumping from the matrix to the intermembrane space (IMS), creating a proton-motive force (Δψ + ΔpH) used by ATP synthase to produce ATP.

Mitchell 1961 — chemiosmotic hypothesis (Nobel 1978) · Boyer & Walker 1997 — rotary mechanism of ATP synthase (Nobel) · MacMunn 1886 — first described cytochromes; Keilin 1925 renamed them · Green — isolated electron transport complexes

Fig. 14.5The electron transport chain and ATP synthase (chemiosmosis). Electrons flow from NADH/FADH₂ → Complex I or II → Ubiquinone (Q) → Complex III → Cytochrome c → Complex IV → O₂. Protons are pumped at Complexes I, III, IV. The resulting H⁺ gradient drives ATP synthesis through F₀F₁-ATP synthase (Complex V). Mitchell chemiosmotic hypothesis, 1961.

**Table 14.5**ATP yield from complete oxidation of one glucose molecule (aerobic, modern values)
Stage	NADH produced	FADH₂ produced	ATP (substrate-level)	ATP (oxidative phos.)
Glycolysis (cytoplasm)	2 NADH	—	2 ATP	2×2.5 = 5 ATP*
Pyruvate oxidation (×2)	2 NADH	—	—	2×2.5 = 5 ATP
Krebs cycle (×2 turns)	6 NADH	2 FADH₂	2 GTP	6×2.5 + 2×1.5 = 18 ATP
Total	10 NADH	2 FADH₂	4 ATP	28 ATP
Grand total (modern)				~30–32 ATP
Old textbook value (P/O: NADH=3, FADH₂=2)				36–38 ATP

*Cytoplasmic NADH from glycolysis must cross the IMM via shuttle systems (malate-aspartate shuttle gives 2.5 ATP; glycerol-3-P shuttle gives 1.5 ATP) — actual yield varies slightly. The figure 30 ATP is the current standard. Some textbooks still state 36–38 ATP; accept either but understand why.

Worked Example — Net ATP Yield from One Glucose

“Calculate the net ATP yield from complete aerobic respiration of one molecule of glucose. Show your working.”

Glycolysis: 2 ATP (substrate-level) + 2 NADH (×2.5 = 5 ATP via ETC). Pyruvate oxidation: 2 NADH (×2.5 = 5 ATP). Krebs cycle (2 turns): 2 GTP + 6 NADH (×2.5 = 15 ATP) + 2 FADH₂ (×1.5 = 3 ATP). Grand total = 2 + 5 + 5 + 2 + 15 + 3 = 32 ATP (or ~30, depending on shuttle assumption). Older value: 36 ATP (P/O = 3 for NADH, 2 for FADH₂).

14.9 Pentose Phosphate Pathway, β-Oxidation & Nitrogen Metabolism

14.9.1 Pentose Phosphate Pathway (PPP)

The hexose monophosphate shunt (PPP) is an alternative route for glucose-6-phosphate oxidation occurring in the cytoplasm (especially in liver, adipose tissue, red blood cells, and adrenal cortex). It does not generate ATP directly. Instead it generates:

NADPH — required for reductive biosynthesis (fatty acid synthesis, cholesterol synthesis, glutathione reduction for antioxidant defence). Glutathione (GSH) protects RBCs from oxidative haemolysis.
Ribose-5-phosphate — required for nucleotide (DNA/RNA) synthesis.

G6PD deficiency (glucose-6-phosphate dehydrogenase deficiency) is the most common enzymopathy in humans (>400 million affected). G6PD catalyses the first step of PPP, producing NADPH. Without NADPH, RBCs cannot regenerate GSH → oxidative stress → haemolysis triggered by drugs (primaquine, dapsone), infections, or fava beans (favism). X-linked recessive; confers some protection against malaria.

Exam Tip: G6PD deficiency → favism is a classic MCQ link. Remember: PPP = NADPH + ribose-5-P (not ATP). NADPH is also required by phagocytes (NADPH oxidase produces reactive oxygen species to kill bacteria) and for cytochrome P450 detoxification reactions in the liver.

14.9.2 β-Oxidation of Fatty Acids

Fatty acids are the most energy-dense fuels (~9 kcal/g vs ~4 kcal/g for carbohydrates). Before oxidation:

FA is activated in the cytoplasm: FA + CoA + ATP → Acyl-CoA + AMP + PPi (acyl-CoA synthetase, thiokinase).
Long-chain acyl-CoA cannot cross the IMM directly — transported as acylcarnitine via the carnitine shuttle (carnitine palmitoyl transferase I & II). CPT-I deficiency causes fatty-acid oxidation disorder.
In the matrix: β-oxidation — repeated 4-step cycle (oxidation, hydration, oxidation, thiolysis) that removes 2 carbons as acetyl-CoA per cycle. Each cycle also yields 1 NADH + 1 FADH₂.

Palmitate (C16:0) calculation: 7 cycles of β-oxidation → 8 acetyl-CoA + 7 NADH + 7 FADH₂. Each acetyl-CoA → ~10 ATP (via Krebs + ETC). 8×10 + 7×2.5 + 7×1.5 = 80 + 17.5 + 10.5 = 108 ATP − 2 (activation cost) = ~106 ATP per palmitate.

14.9.3 Vitamins: Classification, Functions, and Deficiencies

**Table 14.6**Vitamins — fat-soluble and water-soluble
Vitamin	Chemical name	Coenzyme/function	Deficiency disease
Fat-Soluble: A, D, E, K
A	Retinol / retinal	Rhodopsin (visual cycle); epithelial integrity; antioxidant	Night blindness (nyctalopia), xerophthalmia, Bitot’s spots
D	Cholecalciferol (D₃) / ergocalciferol (D₂)	Regulates Ca²⁺ and PO₄ absorption; bone mineralisation; formed in skin by UV	Rickets (children), osteomalacia (adults)
E	Tocopherol	Antioxidant; protects PUFA in membranes; male sterility in rats	Haemolytic anaemia (infants); neurological deficits
K	Phylloquinone (K₁) / menaquinone (K₂)	γ-carboxylation of Glu in clotting factors II, VII, IX, X; protein C, S	Haemorrhagic disease of newborn; prolonged bleeding time
Water-Soluble: B-complex + C
B₁	Thiamine	TPP cofactor for pyruvate dehydrogenase, α-KG DH, transketolase	Beriberi (dry: peripheral neuropathy; wet: cardiac), Wernicke’s encephalopathy
B₂	Riboflavin	FAD, FMN in oxidoreductases (ETC Complex I, II)	Ariboflavinosis: glossitis, angular stomatitis, photophobia
B₃	Niacin (nicotinic acid)	NAD⁺, NADP⁺ — electron carriers in >400 redox reactions	Pellagra (3 Ds: Dermatitis, Diarrhoea, Dementia; 4th D = Death)
B₅	Pantothenic acid	Coenzyme A; acyl carrier protein (ACP) in FA synthesis	Rare; burning-feet syndrome
B₆	Pyridoxine / pyridoxal / pyridoxamine	PLP cofactor for transamination, decarboxylation, glycogen phosphorylase	Peripheral neuropathy; sideroblastic anaemia; convulsions
B₇	Biotin	CO₂ carrier in carboxylases (pyruvate carboxylase, ACC, propionyl-CoA carboxylase)	Dermatitis, alopecia (raw-egg white = avidin binds biotin)
B₉	Folic acid (folate)	THF for one-carbon transfers; purine + dTMP synthesis	Megaloblastic anaemia; neural tube defects (NTDs) in foetus
B₁₂	Cobalamin (cyanocobalamin)	Methionine synthase (methylation); methylmalonyl-CoA mutase	Pernicious anaemia; subacute combined degeneration of cord
C	Ascorbic acid	Antioxidant; cofactor for prolyl and lysyl hydroxylase (collagen); Fe³⁺→Fe²⁺ absorption	Scurvy (Lind 1747; bleeding gums, perifollicular haemorrhage, poor wound healing)

14.9.4 Nitrogen Metabolism and Urea Cycle

Amino groups from protein catabolism are first removed by transamination (transfer of –NH₂ to α-ketoglutarate, forming glutamate) catalysed by aminotransferases (require PLP). Glutamate is then oxidatively deaminated by glutamate dehydrogenase (mitochondrial matrix) releasing NH₃ + α-ketoglutarate. Ammonia is highly toxic to the CNS and must be rapidly detoxified.

The urea cycle (Krebs-Henseleit cycle, 1932) converts toxic NH₃ to non-toxic, water-soluble urea [(NH₂)₂CO] for urinary excretion. It operates partly in the mitochondria (carbamoyl phosphate synthesis, citrulline formation) and partly in the cytoplasm (argininosuccinate synthesis, fumarate release, arginine hydrolysis).

Step 1 (mitochondrial): NH₃ + CO₂ + 2 ATP → carbamoyl phosphate [carbamoyl phosphate synthetase I (CPS-I), rate-limiting; activated by N-acetylglutamate].
Carbamoyl phosphate + ornithine → citrulline (ornithine transcarbamoylase).
Cytoplasmic steps: citrulline + aspartate → argininosuccinate → arginine + fumarate → urea + ornithine (regenerated).
Net: 2 NH₃ + CO₂ → urea + H₂O; costs 3 ATP (4 ~P).

Easy to confuse — α-Keto Acid vs Amino Acid

α-Keto Acid

Has a keto group (C=O) at the α-carbon (adjacent to carboxyl). Examples: pyruvate (α-keto propanoate), oxaloacetate, α-ketoglutarate. These are carbon skeletons after amino-group removal (transamination). They enter the TCA cycle.

Amino Acid

Has an amino group (NH₂) at the α-carbon. Interconverted with α-keto acids by transamination. Amino acids whose carbon skeletons feed into the TCA cycle (glucogenic AAs) or generate acetyl-CoA/acetoacetate (ketogenic AAs) provide energy or gluconeogenic precursors.

Mnemonic — Krebs Cycle Steps

“Citrate Is Krebs’ Starting Substrate For Making Oxaloacetate”

C = Citrate I = Isocitrate K = α-Ketoglutarate S = Succinyl-CoA S = Succinate F = Fumarate M = Malate O = Oxaloacetate

Also: Mnemonic for fat-soluble vitamins — “ADEK” (A, D, E, K). For B-vitamin coenzymes: “The Ribose Niacin Pantothenate Pyridoxine Biotinyl Folyl Cobalamin” = B1–B12 sequence.

14.10 Quick-Reference Tables

**Table 14.7**Regulatory enzymes of glycolysis
Enzyme	Step	Activated by	Inhibited by	Notes
Hexokinase	1	Glucose availability	Glucose-6-phosphate (product inhibition)	High affinity for glucose (low K_m); ubiquitous
Phosphofructokinase-1 (PFK-1)	3	AMP, ADP, fructose-2,6-bisphosphate	ATP, citrate, H⁺	Primary control valve of glycolysis; allosteric; sigmoidal kinetics
Pyruvate kinase	10	Fructose-1,6-bisphosphate (feedforward)	ATP, alanine, acetyl-CoA	Liver isoform (L-PK) also regulated by phosphorylation (glucagon)

**Table 14.8**Essential fatty acids
Fatty acid	Class	C:DB	Why essential	Sources
Linoleic acid	ω-6 PUFA	C18:2 Δ^9,12	No Δ¹²-desaturase in humans	Sunflower, soybean, corn oil
α-Linolenic acid (ALA)	ω-3 PUFA	C18:3 Δ^9,12,15	No Δ¹⁵-desaturase in humans	Flaxseed, walnuts, soybean; HP: sea buckthorn oil
EPA (eicosapentaenoic)	ω-3 PUFA	C20:5	Conditionally essential; converted from ALA (inefficiently)	Fatty fish, fish oil; HP: trout (O. mykiss)
DHA (docosahexaenoic)	ω-3 PUFA	C22:6	Brain, retina; converted from EPA (inefficiently)	Fatty fish, algal oil
Arachidonic acid (AA)	ω-6 PUFA	C20:4	Conditionally essential; prostaglandin precursor	Meat, eggs; converted from linoleic acid

**Table 14.9**Aerobic vs anaerobic ATP yield
Parameter	Aerobic respiration	Lactic fermentation	Ethanol fermentation
Net ATP/glucose	~30–32 ATP	2 ATP	2 ATP
Products	CO₂ + H₂O	2 Lactate	2 Ethanol + 2 CO₂
O₂ required?	Yes	No	No
Organism examples	All aerobes	Animals, Lactobacillus	Yeast (Saccharomyces)
Final electron acceptor	O₂	Pyruvate	Acetaldehyde
Complete glucose oxidation?	Yes	No (partial)	No (partial)

Quick Recap

Carbohydrates: Cn(H₂O)n; monosaccharides (glucose α/β anomers); disaccharides (sucrose = non-reducing; lactose, maltose = reducing); polysaccharides (starch: α-1,4/α-1,6; glycogen: more branched; cellulose: β-1,4; chitin: β-1,4 GlcNAc).
Lipids: FA saturated vs unsaturated; essential FA = linoleic (ω-6) + linolenic (ω-3); trans fats = harmful; triglycerides = energy storage; phospholipids = membrane bilayer.
Proteins: 20 AA; peptide bonds; 4 structural levels; essential AAs = PVT TIM HALL; Pauling 1951 (α-helix & β-sheet); Anfinsen 1972 (folding = primary sequence).
Nucleic acids: DNA (deoxyribose, T, dsDNA, Chargaff: A=T, G≡C); RNA (ribose, U, ssRNA, many types). Nucleotide = base + sugar + phosphate. Purines (A, G): 2 rings; pyrimidines (C, T, U): 1 ring.
Enzymes: protein catalysts; lower E_a; lock-and-key (Fischer 1894) vs induced-fit (Koshland 1958); Michaelis-Menten kinetics; competitive (K_m↑, V_max same) vs non-competitive (V_max↓, K_m same); allosteric + feedback inhibition. EC 1–7 classes.
Glycolysis (EMP): 10 steps, cytoplasm; net 2 ATP + 2 NADH + 2 pyruvate; regulatory enzymes: hexokinase, PFK-1 (primary control), pyruvate kinase.
Pyruvate oxidation: PDC in matrix; pyruvate + CoA + NAD⁺ → acetyl-CoA + NADH + CO₂.
Krebs cycle: 8 steps, matrix; per turn: 3 NADH + 1 FADH₂ + 1 GTP + 2 CO₂; ×2 per glucose.
ETC + oxidative phosphorylation: IMM; Complexes I-IV; Q + Cyt c mobile; H⁺ pumped at I, III, IV; ATP synthase (Complex V); Mitchell chemiosmotic theory (Nobel 1978); net ~30–32 ATP per glucose.
PPP: cytoplasm; produces NADPH + ribose-5-P (no ATP); G6PD deficiency → favism.
β-Oxidation: FA → acetyl-CoA; carnitine shuttle; palmitate → ~106 ATP.
Vitamins: ADEK fat-soluble; B-complex + C water-soluble. Key links: B₁–TPP; B₂–FAD; B₃–NAD; B₅–CoA; B₇–biotin; B₉–folate; B₁₂–cobalamin; C–collagen/antioxidant.
Urea cycle: Krebs-Henseleit 1932; CPS-I (rate-limiting); NH₃ → urea; mitochondrial + cytoplasmic; costs 3 ATP.

Chapter 14 Cheatsheet

Carbohydrates

Reducing sugars: free aldehyde/ketone → Benedict’s/Fehling’s +ve
Sucrose: non-reducing (α,β-1,2); glucose+fructose
Starch: amylose (α-1,4) + amylopectin (α-1,4/α-1,6 every 24-30)
Glycogen: more branched than amylopectin (8-12 residues)
Cellulose: β-1,4 glucose; indigestible by humans; structural
Chitin: β-1,4 GlcNAc; fungal wall + insect exoskeleton

Lipids & Vitamins

Essential FA: linoleic (ω-6) + α-linolenic (ω-3)
Trans fats: partial hydrogenation; raise LDL; harmful
Iodine number ↑ = more unsaturation
ADEK fat-soluble; A = retinol (rhodopsin); D = cholecalciferol (Ca regulation); K = clotting factors
B₃ deficiency = pellagra (3D); B₁ = beriberi; B₉ = NTD; C = scurvy

Enzymes

Lock-key: Fischer 1894; Induced-fit: Koshland 1958
Competitive: K_m↑, V_max same; overcome by ↑[S]
Non-competitive: V_max↓, K_m same; cannot overcome
PFK-1: KEY regulator of glycolysis; allosteric; inhibited by ATP/citrate
Sumner 1926: urease crystallised = protein (Nobel 1946)
EC 2 = Transferase (kinases!); EC 3 = Hydrolase

Glycolysis & Fermentation

10 steps; cytoplasm; net 2 ATP + 2 NADH + 2 pyruvate
Investment: 2 ATP (steps 1, 3); Payoff: 4 ATP + 2 NADH (steps 7, 10)
Regulatory: hexokinase, PFK-1 ★, pyruvate kinase
Anaerobic: lactate (muscle/bacteria) or ethanol (yeast)
HP angle: chhang/lugri (Kinnaur barley beer) = yeast fermentation

Krebs Cycle

Per acetyl-CoA: 3 NADH + 1 FADH₂ + 1 GTP + 2 CO₂
×2 per glucose: 6 NADH + 2 FADH₂ + 2 GTP + 4 CO₂
Citrate synthase ★; isocitrate DH ★
Krebs 1937 (Nobel 1953); Krebs-Henseleit 1932 (urea cycle)

ETC & ATP Yield

Complex I: NADH-Q reductase; pumps 4H⁺
Complex II: no H⁺ pumping (FADH₂ entry)
Complex III + IV: pump 4H⁺ + 2H⁺
Mitchell chemiosmotic theory: Nobel 1978
Net ATP: ~30–32 per glucose (modern); 36–38 (old texts)
NADH ≈ 2.5 ATP; FADH₂ ≈ 1.5 ATP

Proteins & Amino Acids

20 standard AA; L-configuration; zwitterion at pH 7.4
Essential (10): PVT TIM HALL
1° = peptide bond; 2° = H-bonds (α-helix, β-sheet); 3° = all R-group forces; 4° = subunits
Haemoglobin: 2α + 2β (quaternary)
Biuret test = purple (protein); Ninhydrin = purple (free AA)

Discoveries

Wöhler 1828: urea synthesis (end of vitalism)
Fischer 1894: lock-and-key; sugar stereochemistry
Buchner 1897: cell-free fermentation (Nobel 1907)
Sumner 1926: urease = protein (Nobel 1946)
Krebs 1937: TCA cycle (Nobel 1953)
Mitchell 1961: chemiosmosis (Nobel 1978)
Anfinsen 1972: protein folding (Nobel)

Practice Questions

The synthesis of urea from ammonium cyanate (NH₄OCN) by Friedrich Wöhler in 1828 was historically significant because it: HPRCA-pat.

Proved that proteins are polymers of amino acids
Disproved the vitalist doctrine that organic compounds can only be made by living organisms
Demonstrated that enzymes can catalyse inorganic reactions
Established the double-helix structure of DNA

Answer: B — Disproved vitalism

Wöhler synthesised an organic compound (urea) from an inorganic salt in the laboratory, showing that no “vital force” was required. This was the first synthesis of an organic molecule from inorganic precursors and marked the birth of modern organic and biochemistry.

Which disaccharide is non-reducing because both anomeric carbons are involved in the glycosidic bond? HPRCA-pat.

Maltose
Lactose
Sucrose
Cellobiose

Answer: C — Sucrose

Sucrose has an α,β-1,2 glycosidic bond that locks both anomeric carbons (C-1 of glucose and C-2 of fructose), leaving no free reducing group. Maltose and lactose have one free anomeric carbon and are reducing sugars.

The “lock-and-key” hypothesis of enzyme action was proposed by: HPRCA-pat.

Koshland (1958)
Michaelis and Menten (1913)
Emil Fischer (1894)
James Sumner (1926)

Answer: C — Emil Fischer (1894)

Fischer proposed that the enzyme active site has a rigid pre-formed shape complementary to the substrate, like a lock and key. Koshland later proposed the induced-fit modification (1958), showing the enzyme changes shape upon binding.

Which enzyme is considered the primary regulatory (rate-limiting) enzyme of glycolysis?

Hexokinase
Phosphoglucose isomerase
Phosphofructokinase-1 (PFK-1)
Pyruvate kinase

Answer: C — Phosphofructokinase-1 (PFK-1)

PFK-1 catalyses step 3 (fructose-6-P → fructose-1,6-bisphosphate) and is the key allosteric control point. It is activated by AMP/ADP and fructose-2,6-bisphosphate and inhibited by ATP and citrate, making it the primary “valve” of glycolytic flux.

The chemiosmotic theory of ATP synthesis, which earned the Nobel Prize in Chemistry in 1978, was proposed by: HPRCA-pat.

Hans Krebs
Peter Mitchell
Paul Boyer
Fritz Lipmann

Answer: B — Peter Mitchell

Mitchell proposed in 1961 that the proton electrochemical gradient (proton-motive force) across the inner mitochondrial membrane drives ATP synthesis. Boyer and Walker shared the 1997 Nobel for elucidating the rotary mechanism of ATP synthase.

Which of the following vitamins is required as a coenzyme by pyruvate dehydrogenase, α-ketoglutarate dehydrogenase, and transketolase?

Riboflavin (B₂)
Niacin (B₃)
Thiamine (B₁)
Pantothenic acid (B₅)

Answer: C — Thiamine (B₁)

Thiamine pyrophosphate (TPP), the coenzyme form of B₁, is essential for oxidative decarboxylation reactions (PDC, α-KG DH) and for transketolase in the pentose phosphate pathway. Deficiency causes beriberi (affects these pathways in high-energy tissues: nerves, heart).

James Sumner crystallised urease in 1926, earning a share of the Nobel Prize in Chemistry in 1946. The significance of his work was that it demonstrated: HPRCA-pat.

Enzymes are RNA molecules
Enzymes are polysaccharides
Enzymes are proteins
Enzymes require inorganic cofactors for activity

Answer: C — Enzymes are proteins

Until Sumner crystallised urease and showed it was a protein, the chemical nature of enzymes was disputed. His Nobel was shared with Northrop and Stanley (who had crystallised enzymes and viruses, respectively).

Which electron transport chain complex does NOT pump protons across the inner mitochondrial membrane?

Complex I (NADH-Q reductase)
Complex II (Succinate-Q reductase)
Complex III (Q-Cytochrome c reductase)
Complex IV (Cytochrome c oxidase)

Answer: B — Complex II

Complex II (succinate dehydrogenase) accepts electrons from FADH₂ (Krebs cycle) and passes them to ubiquinone but does NOT pump protons. This is why FADH₂ yields fewer ATP than NADH (1.5 vs 2.5 per modern P/O ratio).

Pellagra is caused by deficiency of: HPRCA-pat.

Vitamin B₁ (Thiamine)
Vitamin B₃ (Niacin)
Vitamin B₁₂ (Cobalamin)
Vitamin C (Ascorbic acid)

Answer: B — Vitamin B₃ (Niacin)

Niacin deficiency causes pellagra: the “3 Ds” — Dermatitis (photosensitive skin rash), Diarrhoea, Dementia (and a 4th D: Death if untreated). NAD⁺/NADP⁺ are derived from niacin.

In competitive inhibition of an enzyme, which of the following is TRUE?

V_max decreases; K_m remains unchanged
Both V_max and K_m decrease
V_max remains unchanged; K_m increases
Both V_max and K_m increase

Answer: C — V_max unchanged; K_m increases

In competitive inhibition the inhibitor competes for the active site. Adding more substrate displaces the inhibitor; V_max is eventually reached. But a higher [S] is needed to achieve half V_max, so apparent K_m is elevated. Lineweaver-Burk: lines intersect on the y-axis.

The net ATP yield from the complete aerobic oxidation of ONE molecule of glucose according to modern P/O ratios is approximately: HPRCA-pat.

2 ATP
8 ATP
30–32 ATP
36–38 ATP

Answer: C — 30–32 ATP

Modern P/O ratios (NADH ≈ 2.5, FADH₂ ≈ 1.5) give ~30–32 ATP. Older textbooks used P/O 3 and 2, giving 36–38. HPRCA papers may accept either; mention “modern” value = 30–32.

Which structural feature distinguishes glycogen from amylopectin? HPRCA-pat.

Glycogen contains β-1,4 glycosidic bonds
Glycogen has α-1,6 branches every 8–12 residues (more frequent than amylopectin)
Glycogen contains fructose units
Glycogen is found exclusively in plant cells

Answer: B — More frequent branching in glycogen

Both have α-1,4 main chains and α-1,6 branches, but glycogen branches every 8–12 residues (vs 24–30 in amylopectin). This greater branching allows faster glucose release. Glycogen is the animal/yeast storage form; found in liver and muscle, not plants.

G6PD (glucose-6-phosphate dehydrogenase) deficiency causes haemolytic anaemia upon exposure to primaquine or fava beans because:

There is excess lactate accumulation in RBCs
NADPH production is impaired, preventing regeneration of reduced glutathione
Glycolysis is blocked at step 1
ATP production in RBCs is completely abolished

Answer: B — NADPH deficit → oxidative haemolysis

G6PD catalyses the first step of the PPP generating NADPH. NADPH is essential for reducing glutathione (GSSG → GSH via glutathione reductase). Without GSH, reactive oxygen species destroy RBC membranes. RBCs lack mitochondria and depend entirely on PPP for NADPH.

Which of the following nitrogenous bases is found ONLY in DNA and NOT in RNA? HPRCA-pat.

Adenine
Guanine
Thymine
Cytosine

Answer: C — Thymine

Thymine (5-methyluracil) contains a methyl group at C-5 and is unique to DNA. RNA uses Uracil instead. Adenine, Guanine, and Cytosine are found in both DNA and RNA.

Aspirin (acetylsalicylic acid) is an irreversible inhibitor of cyclooxygenase (COX). This is an example of: HPRCA-pat.

Competitive inhibition
Uncompetitive inhibition
Non-competitive inhibition
Irreversible (covalent) inhibition

Answer: D — Irreversible inhibition

Aspirin acetylates a serine residue in the active site of COX-1 and COX-2 — a covalent modification that permanently inactivates the enzyme until new enzyme is synthesised. This is why the antiplatelet effect of low-dose aspirin lasts the lifetime of the platelet (~7–10 days).

Assertion (A): Sucrose is a non-reducing sugar.
Reason (R): In sucrose, the anomeric carbon of glucose and the anomeric carbon of fructose are both involved in the glycosidic bond.

Both A and R are true and R is the correct explanation of A
Both A and R are true but R is not the correct explanation of A
A is true but R is false
A is false but R is true

Answer: A — Both true; R explains A

Sucrose has an α(1→2)β glycosidic bond linking C-1 of glucose and C-2 of fructose. Both anomeric carbons are blocked, leaving no free reducing group. Hence sucrose cannot reduce copper in Benedict’s/Fehling’s tests.

Assertion (A): The α-helix and β-pleated sheet are both examples of secondary protein structure.
Reason (R): Both are stabilised by hydrogen bonds between the R groups of amino acid side chains.

Both A and R are true and R is the correct explanation of A
Both A and R are true but R is not the correct explanation of A
A is true but R is false
A is false but R is true

Answer: C — A true, R false

Both structures are secondary protein structure (A true). However, they are stabilised by H-bonds between the backbone carbonyl (C=O) and amino (N–H) groups — NOT the R groups (R is false). R-group interactions determine tertiary structure.

Assertion (A): FADH₂ donates electrons to Complex II (succinate dehydrogenase) which does not pump protons, producing fewer ATP than NADH.
Reason (R): The number of ATP produced by a reduced coenzyme depends on how many proton-pumping complexes its electrons pass through.

Both A and R are true and R is the correct explanation of A
Both A and R are true but R is not the correct explanation of A
A is true but R is false
A is false but R is true

Answer: A — Both true; R explains A

FADH₂ feeds electrons into Q directly through Complex II (no proton pumping), bypassing Complex I. Electrons then pass Complexes III and IV only. NADH electrons pass Complexes I, III, and IV. Fewer protons pumped = less proton-motive force = fewer ATP. Modern P/O: NADH = 2.5, FADH₂ = 1.5.

Assertion (A): Feedback inhibition prevents overproduction of end products in metabolic pathways.
Reason (R): In feedback inhibition, the end product binds the active site of the first enzyme in the pathway, blocking substrate entry.

Both A and R are true and R is the correct explanation of A
Both A and R are true but R is not the correct explanation of A
A is true but R is false
A is false but R is true

Answer: C — A true, R false

Feedback inhibition does prevent overproduction (A true). However, end-product inhibition is allosteric — the inhibitor binds a separate allosteric site, not the active site (R false). The conformational change at the allosteric site reduces catalytic activity.

Assertion (A): Cells that lack mitochondria (such as mature mammalian erythrocytes) cannot perform oxidative phosphorylation.
Reason (R): The electron transport chain and ATP synthase are located on the inner mitochondrial membrane.

Both A and R are true and R is the correct explanation of A
Both A and R are true but R is not the correct explanation of A
A is true but R is false
A is false but R is true

Answer: A — Both true; R explains A

Mature RBCs lack nuclei AND mitochondria. Without the inner mitochondrial membrane (the site of ETC and ATP synthase), oxidative phosphorylation is impossible. RBCs rely entirely on glycolysis for ATP and on the pentose phosphate pathway for NADPH.

Match the vitamin with its deficiency disease: HPRCA-pat.

Column I (Vitamin)	Column II (Deficiency disease)
(a) Vitamin A	(i) Pellagra
(b) Vitamin B₁	(ii) Scurvy
(c) Vitamin B₃	(iii) Night blindness
(d) Vitamin C	(iv) Beriberi

a-iii, b-iv, c-i, d-ii
a-iv, b-iii, c-ii, d-i
a-i, b-ii, c-iii, d-iv
a-iii, b-i, c-iv, d-ii

Answer: A — a-iii, b-iv, c-i, d-ii

Vitamin A deficiency → night blindness/xerophthalmia; B₁ deficiency → beriberi; B₃ deficiency → pellagra (3Ds); C deficiency → scurvy (Lind 1747).

Match the biochemist with their discovery: HPRCA-pat.

Column I (Scientist)	Column II (Discovery)
(a) Buchner	(i) TCA/citric acid cycle
(b) Krebs	(ii) Urease is a protein; first enzyme crystallised
(c) Sumner	(iii) Chemiosmotic theory of ATP synthesis
(d) Mitchell	(iv) Cell-free fermentation

a-iv, b-i, c-ii, d-iii
a-i, b-iv, c-iii, d-ii
a-ii, b-i, c-iv, d-iii
a-iv, b-ii, c-i, d-iii

Answer: A — a-iv, b-i, c-ii, d-iii

Buchner 1897 (cell-free fermentation, Nobel 1907); Krebs 1937 (TCA cycle, Nobel 1953); Sumner 1926 (urease crystallised = protein, Nobel 1946); Mitchell 1961 (chemiosmotic theory, Nobel 1978).

Match the enzyme class (IUBMB) with its example:

Column I (Class)	Column II (Example)
(a) Oxidoreductase	(i) Hexokinase
(b) Transferase	(ii) Amylase
(c) Hydrolase	(iii) Lactate dehydrogenase
(d) Lyase	(iv) Aldolase

a-iii, b-i, c-ii, d-iv
a-i, b-ii, c-iv, d-iii
a-iii, b-iv, c-ii, d-i
a-ii, b-iii, c-i, d-iv

Answer: A — a-iii, b-i, c-ii, d-iv

LDH is an oxidoreductase (EC 1, interconverts pyruvate/lactate with NAD⁺/NADH); hexokinase is a transferase (EC 2, transfers phosphoryl group); amylase is a hydrolase (EC 3, cleaves starch by hydrolysis); aldolase is a lyase (EC 4, cleaves fructose-1,6-BP without water or oxidation).

Match the polysaccharide with its correct linkage and organism: HPRCA-pat.

Column I	Column II
(a) Cellulose	(i) α-1,4 + α-1,6; animal/yeast
(b) Chitin	(ii) β-1,4; plant cell wall
(c) Glycogen	(iii) α-1,4 + α-1,6; plant storage
(d) Amylopectin	(iv) β-1,4 GlcNAc; fungi/arthropods

a-ii, b-iv, c-i, d-iii
a-iv, b-ii, c-iii, d-i
a-iii, b-i, c-iv, d-ii
a-ii, b-i, c-iv, d-iii

Answer: A — a-ii, b-iv, c-i, d-iii

Cellulose: β-1,4 glucose (plant cell wall); chitin: β-1,4 N-acetylglucosamine (fungal wall, insect exoskeleton); glycogen: α-1,4/α-1,6 (animal/yeast, more branched); amylopectin: α-1,4/α-1,6 (plant starch component, less branched).

Consider the following statements about the Krebs (TCA) cycle:

It occurs in the mitochondrial matrix.
One turn of the cycle produces 3 NADH, 1 FADH₂, 1 GTP, and 2 CO₂.
Citrate synthase is the rate-limiting regulatory enzyme of the cycle.
Two turns of the cycle are required to completely oxidise one molecule of glucose.

Which of the above are correct?

I and II only
I, II and IV only
I, II, III and IV
II and III only

Answer: C — I, II, III and IV (all four)

All statements are correct. The TCA cycle runs in the mitochondrial matrix; one turn (per acetyl-CoA) gives 3 NADH + 1 FADH₂ + 1 GTP + 2 CO₂; citrate synthase is a key regulatory enzyme; and since one glucose generates 2 pyruvates (2 acetyl-CoA), 2 turns of the cycle are needed per glucose.

Consider the following statements about protein structure:

The primary structure refers to the linear sequence of amino acids linked by peptide bonds.
The α-helix is stabilised by hydrogen bonds between R groups of amino acids.
Disulphide bonds (Cys–Cys) contribute to tertiary structure.
Haemoglobin is an example of a protein with quaternary structure.

Which are correct?

I and II only
I, III and IV only
II and IV only
I, II, III and IV

Answer: B — I, III and IV only

Statement II is false: α-helix H-bonds are between backbone C=O and N–H groups (not R groups). Statements I, III, and IV are correct. R-group interactions (hydrophobic, ionic, H-bond) and disulphide bonds govern tertiary structure. Haemoglobin (2α + 2β) has quaternary structure.

Consider the following statements about the pentose phosphate pathway (PPP):

It occurs in the cytoplasm.
The primary products are NADPH and ribose-5-phosphate.
It generates 2 ATP per glucose (net) by substrate-level phosphorylation.
G6PD deficiency impairs PPP, reducing NADPH availability in red blood cells.

Which are correct?

I, II and IV only
I, II, III and IV
II and III only
I and IV only

Answer: A — I, II and IV only

Statement III is false: PPP does NOT directly generate ATP. Its main outputs are NADPH (for reductive biosynthesis and antioxidant defence) and ribose-5-phosphate (for nucleotide synthesis). Statements I, II, and IV are correct.

Arrange the following biochemistry discoveries in correct chronological order: HPRCA-pat.

Wöhler — synthesis of urea (end of vitalism)
Buchner — cell-free fermentation
Krebs — TCA cycle
Mitchell — chemiosmotic theory

I → II → III → IV
II → I → IV → III
I → III → II → IV
III → I → II → IV

Answer: A — I → II → III → IV

Wöhler 1828 → Buchner 1897 → Krebs 1937 → Mitchell 1961. These are four landmark dates in biochemistry that are reliably tested; memorise the timeline.

Which of the following is the odd one out in terms of its role in the electron transport chain?

Complex I (NADH-Q reductase)
Complex III (Q-cyt c reductase)
Complex IV (Cyt c oxidase)
Complex II (Succinate-Q reductase)

Answer: D — Complex II

Complexes I, III, and IV all pump protons across the inner mitochondrial membrane, contributing to the proton-motive force for ATP synthesis. Complex II does not pump protons — it is the odd one out. This also explains why FADH₂ (entering via Complex II) produces fewer ATP than NADH.

End of Chapter 14 · Biochemistry. HPRCA-pat. indicates HPRCA / state-TGT pattern questions; literal past-paper items will be flagged with year when official papers are sourced.

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