(a) (i) Explain the significance of Nod factors and the process of root nodule formation. (10 marks) (ii) Describe the process…

(a)(i) Significance of Nod factors and the process of root nodule formation (10 marks)

Nod factors are the signal molecules produced by rhizobia that trigger nodulation in legumes. Chemically they are lipo-chitooligosaccharides: a backbone of three to five beta-1,4-linked N-acetylglucosamine units with a fatty acyl chain on the non-reducing end, and species-specific decorations (sulphate, acetyl, methyl or fucosyl groups). They are made by the products of the nod genes: nodA, nodB and nodC are common (nodC is a chitin synthase), while host-specific genes add the decorations, and nodD, activated by flavonoids exuded by the legume root (luteolin, daidzein, genistein), switches the nod genes on.

Significance:

• They determine host specificity, as the plant recognises only the correct decorations through LysM receptor kinases (NFR1 and NFR5 in Lotus).
• They induce root hair curling and deformation, calcium spiking, expression of early nodulin genes, cortical cell division and infection thread formation, and so are sufficient to start nodule development.

Process of nodule formation:

1. Flavonoids from the root attract rhizobia and activate nodD.
2. Bacteria attach to root hairs, and the Nod factors curl the hair (shepherd's crook).
3. An infection thread forms by invagination and carries the dividing bacteria to the cortex.
4. Cortical cells divide to form the nodule primordium, from the inner cortex in soybean (determinate nodule) or outer cortex in pea (indeterminate).
5. Bacteria are released into the cells by endocytosis, enclosed by a symbiosome membrane, and become nitrogen-fixing bacteroids.
6. Leghaemoglobin (pink colour) keeps oxygen low, nitrogenase reduces N₂ to ammonia, and the plant supplies malate while receiving fixed nitrogen.

(a)(ii) Amino acid biosynthesis in nitrogen metabolism (10 marks)

Amino acids are made from assimilated nitrogen and carbon skeletons from glycolysis, the TCA cycle and the pentose phosphate pathway.

Nitrogen assimilation. Nitrate is reduced to nitrite by nitrate reductase (cytosol, NADH) and to ammonium by nitrite reductase (plastids, reduced ferredoxin). Ammonium, from nitrate, nitrogen fixation or photorespiration, is assimilated mainly by the GS-GOGAT cycle:

• Glutamine synthetase: glutamate + NH₄⁺ + ATP → glutamine.
• Glutamate synthase (GOGAT): glutamine + 2-oxoglutarate + reduced ferredoxin or NADH → two glutamate.
• Glutamate dehydrogenase can also aminate 2-oxoglutarate to glutamate when ammonium is high.

Formation of other amino acids:

• Transamination: aminotransferases with pyridoxal phosphate move the amino group from glutamate to an oxo acid; aspartate aminotransferase makes aspartate from oxaloacetate, and alanine aminotransferase makes alanine from pyruvate.
• Amidation: asparagine synthetase forms asparagine from aspartate and glutamine.
• Families derived from a common precursor: glutamate (Gln, Pro, Arg), aspartate (Asn, Lys, Met, Thr, Ile), pyruvate (Ala, Val, Leu), 3-phosphoglycerate (Ser, Gly, Cys), and the shikimate pathway (Phe, Tyr, Trp), with histidine from ribose phosphate.

Glutamine and glutamate are the main donors of nitrogen for all other nitrogenous compounds.

(b) Photophosphorylation versus oxidative phosphorylation (15 marks)

Both make ATP by chemiosmosis (Mitchell), with an electron transport chain that builds a proton gradient and an ATP synthase driven by that gradient. They differ as follows:

• Site: photophosphorylation is in the thylakoid membranes of chloroplasts, and oxidative phosphorylation in the inner mitochondrial membrane.
• Energy source: light absorbed by photosystems in the first, and the energy released by oxidation of NADH and FADH₂ in the second.
• Electron donor: water in photophosphorylation (releasing O₂), and NADH and FADH₂ in oxidative phosphorylation.
• Final electron acceptor: NADP⁺ in non-cyclic photophosphorylation, and oxygen (forming water) in oxidative phosphorylation.
• Carriers: photosystem II, plastoquinone, cytochrome b6f, plastocyanin, photosystem I and ferredoxin, against complexes I to IV, ubiquinone and cytochrome c.
• Direction of electron flow: uphill, against the redox gradient, driven by light, in the Z-scheme, and downhill, exergonic, from NADH to O₂.
• Proton gradient: protons are pumped into the thylakoid lumen, and ATP is made on the stroma side by the CF₀-CF₁ ATP synthase; in mitochondria they are pumped into the intermembrane space and ATP is made in the matrix by F₀-F₁.
• Types: photophosphorylation may be cyclic (only ATP, involving photosystem I) or non-cyclic (ATP, NADPH and O₂); oxidative phosphorylation has one type.
• Occurrence: only in light, in photosynthetic cells; oxidative phosphorylation occurs in the light and in the dark, in all aerobic cells.
• Use: the ATP of photophosphorylation goes into the Calvin cycle, and that of oxidative phosphorylation into cellular work.

(c) Biochemical and physiological changes during fruit ripening (15 marks)

Fruit ripening is the change of a mature fruit into an edible one, in which the fruit softens, changes colour, becomes sweet and aromatic. Climacteric fruits (apple, banana, tomato, mango) show a burst of respiration and an autocatalytic rise of ethylene, while non-climacteric fruits (citrus, grape, strawberry) do not.

Physiological changes:

• A respiratory climacteric in climacteric fruits, and a surge of ethylene made by ACC synthase and ACC oxidase, acting through its receptor (ETR1) and the signalling pathway CTR1, EIN2 and EIN3.
• Increased membrane permeability, loss of compartmentation and a rise in ripening-specific gene expression under transcription factors such as RIN and NOR in tomato.

Biochemical changes:

• Softening: cell wall-degrading enzymes (polygalacturonase, pectin methylesterase, cellulase, expansin) break down protopectin into soluble pectin, and the middle lamella loosens.
• Colour: chlorophyll breaks down, and carotenoids (lycopene in tomato) and anthocyanins are synthesised as chloroplasts become chromoplasts.
• Sweetness: starch is hydrolysed to sugars (amylase and phosphorylase), and organic acids fall as they are used in respiration.
• Flavour: volatile esters, alcohols, aldehydes and terpenes form (isoamyl acetate in banana).
• Astringency falls, as tannins polymerise, and toxic alkaloids (solanine) decline.
• Protein synthesis and hormone changes occur, with ABA and ethylene rising.

Control: ethylene (or ethephon) is used to ripen fruits, while 1-MCP, low temperature, low O₂ with high CO₂, and calcium slow ripening in storage.