(a) Draw the molecular structure of ATP synthase and diagrammatically explain the chemiosmotic concept of ATP synthesis. 20 (b)…

(a) ATP synthase and the chemiosmotic concept of ATP synthesis

ATP synthase (F₁F₀-ATPase, complex V) is a rotary enzyme of the inner mitochondrial membrane. The labelled diagram of its molecular structure should show:

• F₀ in the membrane: subunit a with two half-channels for H⁺, two b subunits, and a ring of c subunits (eight in vertebrates)
• F₁ projecting into the matrix: an α₃β₃ hexamer with the catalytic sites on the three β subunits
• the central stalk (γ, ε) joining the c-ring to the hexamer, and the peripheral stalk (b, OSCP) holding the hexamer still

Chemiosmotic concept (Peter Mitchell, 1961; Nobel Prize 1978). The second diagram should show these steps, the intermembrane space H⁺-rich (positive) and the matrix H⁺-poor (negative):

1. Complexes I, III and IV of the electron transport chain pump H⁺ from the matrix into the intermembrane space (about 10 H⁺ per NADH) as electrons pass to O₂.
2. The impermeable inner membrane holds the gradient as the proton-motive force, Δp = Δψ − (2.303RT/F)ΔpH (inside minus outside), about 0.2 V in magnitude (Δψ about 0.14 V plus about 60 mV per pH unit).
3. H⁺ flows back through F₀ down this gradient; protonation and deprotonation of carboxyl groups on the c subunits turn the c-ring and the γ stalk.
4. The turning γ drives the three β subunits through open, loose and tight states (Boyer's binding-change mechanism): ADP and Pi bind in the loose state, ATP forms in the tight state and is released in the open state. One 360° turn makes 3 ATP.
5. With a c₈ ring, 8 H⁺ per turn give 2.7 H⁺ per ATP, about 3.7 including H⁺ for Pi and ADP/ATP transport.

Evidence: uncouplers such as 2,4-dinitrophenol dissipate the gradient, so electron transport continues but ATP synthesis stops; oligomycin blocks F₀; an artificial pH gradient made ATP in chloroplasts (Jagendorf, 1966) and in vesicles with bacteriorhodopsin and ATP synthase (Racker and Stoeckenius, 1974).

(b)(i) A=T and G≡C base pairing in the DNA double strand

The labelled diagram should show:

• two antiparallel sugar–phosphate strands, one running 5′→3′ and the other 3′→5′
• adenine (a purine) joined to thymine (a pyrimidine) by two hydrogen bonds, A=T
• guanine (a purine) joined to cytosine (a pyrimidine) by three hydrogen bonds, G≡C
• base pairs stacked 0.34 nm apart, in a helix 2 nm wide with one turn of 3.4 nm per 10 base pairs

Pairing is complementary (Watson and Crick, 1953). A purine always pairs with a pyrimidine, which keeps the width of the helix uniform, and the hydrogen-bonding groups allow only A with T and G with C, so Chargaff's rule (A = T, G = C) holds. The extra bond makes G≡C pairs stronger, so GC-rich DNA melts at a higher temperature.

(b)(ii) Michaelis–Menten kinetic pattern of an enzymatic reaction

For E + S ⇌ ES → E + P (Michaelis and Menten, 1913; steady-state treatment by Briggs and Haldane, 1925), the initial velocity is v = Vmax[S]/(Km + [S]), where Km = (k₋₁ + k₂)/k₁ and Vmax = k₂[E]total. The labelled diagram should show:

• a rectangular hyperbola of v against [S]: nearly straight at low [S] (first order), bending over to approach Vmax at high [S] (zero order, enzyme saturated)
• Vmax as the plateau, and Km as the [S] at which v = Vmax/2
• a Lineweaver–Burk plot of 1/v against 1/[S]: a straight line with y-intercept 1/Vmax, x-intercept −1/Km and slope Km/Vmax

A low Km means high affinity for the substrate.

(c) Cellular mechanism of action of steroid hormones

Steroid hormones (cortisol, aldosterone, oestradiol, progesterone, testosterone) are lipid-soluble derivatives of cholesterol. They travel in the blood bound to carrier proteins and diffuse through the plasma membrane of target cells, so they act on intracellular receptors, unlike peptide hormones.

Genomic action:

1. The inactive receptor lies in the cytoplasm (glucocorticoid, mineralocorticoid) or already in the nucleus (oestrogen, progesterone), held by heat-shock proteins such as hsp90.
2. The hormone binds the ligand-binding domain; the receptor changes shape and releases hsp90.
3. The hormone–receptor complex dimerises and, if cytoplasmic, enters the nucleus through the nuclear pore.
4. Its DNA-binding domain, with two zinc-containing motifs, binds the hormone response element (for example the glucocorticoid response element) in the promoter or enhancer of target genes.
5. The complex recruits coactivators or corepressors and chromatin-modifying enzymes, so that RNA polymerase II transcribes or represses the gene.
6. The mRNA is translated into new proteins (for example gluconeogenic enzymes in the liver under cortisol), which produce the cellular response.

The response is slow (hours) but long-lasting and amplified, because each gene gives many mRNAs and proteins. Some steroid effects occur within seconds to minutes through membrane-associated receptors and second messengers (non-genomic action). The signal ends when the hormone dissociates and is metabolised.