Metabolic Pathways
7 sub-topics · Pages 210–271
1. Introduction
Microbial central metabolism — glycolysis, the TCA cycle, and oxidative phosphorylation — converts diverse organic substrates into ATP and biosynthetic building blocks. These pathways are amphibolic: simultaneously catabolic (energy-releasing) and anabolic (supplying precursors for biosynthesis). Understanding central metabolism is essential for designing bioremediation strategies, optimising fermentation processes, and identifying antibiotic targets.
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1. The anaerobic breakdown of glucose to pyruvate in the cytoplasm, yielding 2 ATP and 2 NADH, is called _______.
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Glycolysis🔘 Multiple Choice
1. The maximum ATP yield per glucose molecule is highest in which process?
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Correct: C) Aerobic respiration (~30–32 ATP)2. In the pentose phosphate pathway (PPP), the main products used by the cell are:
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Correct: B) NADPH (for biosynthesis and defence against oxidative stress) and pentose sugars (for nucleotide synthesis)3. Which pigment is the primary photosynthetic pigment in cyanobacteria?
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Correct: C) Chlorophyll a💬 Open-Ended Questions
1. Explain how photoautotrophic microorganisms (e.g., Cyanobacteria) use light energy to power both carbon fixation and nitrogen fixation simultaneously. What are the energy requirements of each process?
Hint / Guidance
Light reactions: photosystems I and II generate ATP and NADPH from light + H₂O (PS II: H₂O → O₂ + e⁻; PS I: ferredoxin → NADPH); also generate reduced ferredoxin. Carbon fixation (Calvin cycle): 3 CO₂ + 9 ATP + 6 NADPH → 1 G3P. N₂ fixation: N₂ + 16 ATP + 8 e⁻ + 8 H⁺ → 2 NH₃ + H₂ (nitrogenase reaction); extremely energy-expensive. Simultaneous: in vegetative cells (photosynthesis); in heterocysts (N₂ fixation — Photosystem II absent, O₂ protection). ATP and reductant from photosynthesis power both processes.2. How do heterocyst-forming cyanobacteria (e.g., Anabaena) solve the problem of protecting nitrogenase from oxygen while still performing oxygenic photosynthesis?
Hint / Guidance
Heterocysts: ~5–10% of cells differentiate into thick-walled, non-dividing cells; lose PSII (no O₂ evolution); thick glycolipid envelope restricts O₂ diffusion; high respiration rate consumes residual O₂; produce ATP/reductant via PSI + heterocyst-specific ferredoxin for nitrogenase. Vegetative cells fix CO₂ and send sucrose to heterocysts; heterocysts send glutamine (fixed N) to vegetative cells. Spatial separation of O₂-producing and N₂-fixing reactions.2. Catabolic Diversity
Microorganisms exhibit extraordinary catabolic diversity, exploiting energy sources ranging from glucose and amino acids to cellulose, petroleum hydrocarbons, and inorganic compounds. Aerobic heterotrophs, anaerobic fermenters, sulfate reducers, methanogens, and chemolithotrophs each occupy distinct ecological niches defined by their energy metabolism. This metabolic breadth makes microbes the primary drivers of biogeochemical cycling on Earth.
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1. In anaerobic respiration, bacteria can use alternative final electron acceptors such as nitrate (NO₃⁻) or sulfate (SO₄²⁻) instead of _______.
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Oxygen (O₂)🔘 Multiple Choice
1. In glycolysis, the 6-carbon glucose molecule is split into two 3-carbon molecules of:
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Correct: C) Pyruvate2. Nitrification is the two-step conversion of:
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Correct: B) NH₄⁺ → NO₂⁻ → NO₃⁻💬 Open-Ended Questions
1. Compare aerobic respiration, anaerobic respiration, and fermentation. For each: identify the final electron acceptor, approximate ATP yield per glucose, and give an example organism and environment.
Hint / Guidance
Aerobic: O₂, ~30–32 ATP, E. coli (oxic gut). Anaerobic respiration: NO₃⁻/SO₄²⁻/Fe³⁺ etc., ~17–19 ATP (nitrate), ~11 ATP (sulfate), Desulfovibrio in anoxic sediment. Fermentation: organic molecule (e.g., pyruvate or acetaldehyde), 2 ATP, Lactobacillus (lactic acid in yogurt), Saccharomyces (ethanol in beer). Comparison: aerobic most efficient; fermentation least; anaerobic respiration intermediate; all use glycolysis.2. A wastewater treatment plant relies on aerobic heterotrophic bacteria to remove BOD. A power failure shuts down the aeration system for 6 hours. Describe what happens metabolically to the microbial community and what the effects are on treatment efficiency.
Hint / Guidance
Without O₂: facultative anaerobes switch to anaerobic respiration (using NO₃⁻ if available) then fermentation; obligate aerobes stop growing and begin dying; NADH accumulates; organic acids and alcohols produced by fermentation lower pH; nitrification (aerobic, strict aerobes Nitrosomonas/Nitrobacter) completely stops; BOD removal drops dramatically; effluent quality deteriorates (high BOD, ammonia). After restoration: aerobic populations recover (lag phase); complete recovery may take 24–48 h; risk of bulking if floc structure disrupted.3. Explain how the Wood-Ljungdahl (reductive acetyl-CoA) pathway functions and which organisms use it. What is its significance for the early evolution of metabolism?
Hint / Guidance
WL pathway: CO₂ reduced to CO (carbonyl branch via CO dehydrogenase) + CO₂ reduced to methyl-THF (methyl branch); combined by CODH/ACS to form acetyl-CoA; used by acetogens (Acetobacterium) and methanogens (Methanobacterium). Only pathway that can serve as both CO₂ fixation route AND energy-conservation pathway. Evolutionary significance: proposed as first metabolic pathway at origin of life; operates in anaerobic, H₂-rich conditions similar to early Earth; runs in reverse as energy source (acetogenesis).3. Krebs Cycle
The TCA cycle (Krebs cycle) oxidises acetyl-CoA to 2CO₂, generating 3 NADH, 1 FADH₂, and 1 GTP per turn. These electron carriers feed the electron transport chain, ultimately driving the synthesis of ~10 ATP per acetyl-CoA via chemiosmosis. The cycle is also amphibolic: intermediates (oxaloacetate, α-ketoglutarate, succinyl-CoA) are biosynthetic precursors for amino acids, nucleotides, and porphyrins.
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1. The cyclic pathway in which acetyl-CoA is oxidised to CO₂ while generating NADH, FADH₂, and GTP is called the _______ cycle.
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Tricarboxylic acid (TCA / Krebs)🔘 Multiple Choice
1. The function of NAD⁺ regeneration in fermentation is to:
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Correct: B) Allow glycolysis to continue by re-oxidising NADH to NAD⁺ so the glycolytic pathway is not blocked2. The enzyme nitrogenase, which catalyses nitrogen fixation, is irreversibly inhibited by:
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Correct: C) Oxygen💬 Open-Ended Questions
1. Describe the electron transport chain in aerobic bacteria. Explain how the proton motive force is generated and how it drives ATP synthesis via chemiosmosis.
Hint / Guidance
NADH and FADH₂ (from glycolysis/TCA) donate electrons to Complex I (NADH dehydrogenase) or Complex II (succinate dehydrogenase). Electrons pass through ubiquinone to Complex III (bc₁), then cytochrome c to Complex IV (cytochrome c oxidase). O₂ is the terminal acceptor → H₂O. Protons pumped outward at Complexes I, III, IV → PMF (ΔpH + Δψ). Protons flow back through ATP synthase (F₀F₁) → conformational change drives ADP + Pᵢ → ATP (chemiosmosis, Mitchell's hypothesis).2. Describe the β-oxidation pathway for fatty acid catabolism in bacteria. Why do organisms that can catabolise fatty acids have a significant competitive advantage in certain environments?
Hint / Guidance
β-oxidation: fatty acid + CoA → fatty acyl-CoA (activated); successive cycles of oxidation at β-carbon: FAD-linked dehydrogenation → FADH₂; hydration; NAD⁺-linked dehydrogenation → NADH; thiolysis releases acetyl-CoA and shortened fatty acyl-CoA. Each 2-carbon unit yields: 1 acetyl-CoA + 1 NADH + 1 FADH₂ → fed into TCA + ETC. Energy yield: palmitate (16C) → 7 cycles + 8 acetyl-CoA → ~108 ATP. Competitive advantage: in environments with lipid-rich substrates (hydrocarbon spills, oil seeps, adipose tissue infection); fatty acids provide higher ATP/carbon than carbohydrates; organisms like Pseudomonas can degrade petroleum hydrocarbons.3. What is the significance of the anammox (anaerobic ammonium oxidation) process in the nitrogen cycle? Name the organisms involved and describe the unique cellular feature associated with this pathway.
Hint / Guidance
Anammox: NH₄⁺ + NO₂⁻ → N₂ + 2H₂O; removes fixed nitrogen from ecosystems; responsible for ~50% of N₂ production in marine systems; important in wastewater treatment (SHARON-ANAMMOX process). Organisms: Candidatus Brocadia, Kuenenia, Scalindua (all Planctomycetes). Unique feature: anammoxosome — specialised membrane-bound compartment where hydrazine (N₂H₄) is an intermediate; membrane contains unique ladderane lipids (concatenated cyclobutane rings) that reduce permeability to the toxic intermediate hydrazine.4. Methanogenesis
Methanogenesis is carried out exclusively by methanogenic Archaea — strict anaerobes found in anoxic sediments, ruminant guts, and anaerobic digesters. They reduce CO₂ with H₂ or cleave acetate to produce methane (CH₄). Methane is both a potent greenhouse gas (28× CO₂ over 100 years) and a renewable energy source. Methanogenesis is the terminal step in anaerobic decomposition, driving the complete mineralisation of organic matter in oxygen-free environments.
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1. Microorganisms that obtain energy by oxidising inorganic compounds such as ammonia, hydrogen sulfide, or ferrous iron are called _______.
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Chemolithotrophs🔘 Multiple Choice
1. Which statement about the electron transport chain (ETC) is correct?
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Correct: B) The ETC transfers electrons through a series of carriers, pumping protons across the membrane to generate a proton motive force that drives ATP synthase2. Anoxygenic photosynthesis differs from oxygenic photosynthesis in that it:
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Correct: B) Uses H₂S or organic compounds as electron donors and does not produce O₂💬 Open-Ended Questions
1. Sulfate-reducing bacteria (SRB) cause significant problems in engineered systems. Describe their metabolism, and explain how they contribute to (a) biocorrosion of metal infrastructure and (b) souring of oil reservoirs.
Hint / Guidance
SRB (e.g., Desulfovibrio) use SO₄²⁻ as terminal electron acceptor → H₂S + oxidised organic compounds. (a) Biocorrosion: H₂S reacts with iron → FeS (pitting corrosion); SRB consume cathodic H₂ from iron surface (cathodic depolarisation) accelerating iron oxidation; biofilms create localised anaerobic micro-environments. (b) Souring: injected seawater brings SO₄²⁻ into oil reservoir; SRB oxidise petroleum hydrocarbons using SO₄²⁻ → H₂S contaminates oil/gas; reduces product value, corrodes pipelines, creates H₂S safety hazard.2. Explain the mixed-acid fermentation pathway in E. coli. What products are formed and under what conditions? How is the methyl red test used to detect this fermentation?
Hint / Guidance
Mixed-acid fermentation (E. coli at low pH or under stress): pyruvate → formate + acetyl-CoA (pyruvate formate lyase); formate → CO₂ + H₂ (formate hydrogen lyase); acetyl-CoA → ethanol or acetate; OAA → succinate. Products: formic acid, acetic acid, ethanol, lactic acid, CO₂, H₂, succinic acid — mixture of acids significantly lowers pH. Methyl red test: cultures grown in glucose-phosphate medium; acidic end-products → pH < 4.4 → methyl red indicator turns red (MR+, e.g., E. coli). Contrast with Voges-Proskauer test (acetoin/2,3-butanediol, neutral products, e.g., Klebsiella).3. Describe reverse electron transport and explain why some chemolithotrophic bacteria must use it. Give a specific example.
Hint / Guidance
Reverse electron transport: electrons from low-reduction-potential donors (e.g., Fe²⁺, E₀' = +0.77V; NO₂⁻, E₀' = +0.43V) enter ETC at ubiquinol level; they cannot reduce NAD⁺ (E₀' = -0.32V) spontaneously; instead, PMF drives electrons backward through Complex I to reduce NAD⁺, consuming ATP. Example: Nitrobacter winogradskyi oxidises NO₂⁻ → NO₃⁻ via nitrite oxidoreductase (NXR); electrons enter at ubiquinol; some flow forward to O₂ for ATP via Complex IV; some driven backward to generate NADH for CO₂ fixation via Calvin cycle.5. Krebs Cycle
Anaplerotic reactions replenish TCA cycle intermediates that are continuously drained for biosynthesis. The most important is pyruvate carboxylase (pyruvate + CO₂ → oxaloacetate), which replenishes oxaloacetate when it is consumed in gluconeogenesis. Without anaplerosis, the TCA cycle would grind to a halt as intermediates were depleted — making these reactions essential even when the cycle is running at full capacity.
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1. The metabolic process in which organic substrates are incompletely oxidised using an organic molecule as the final electron acceptor (no external terminal acceptor), generating only 2 ATP per glucose, is called _______.
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Fermentation2. In the light reactions of photosynthesis, water is split and O₂ is released in a process called _______.
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Photolysis (water splitting / photolysis of water)🔘 Multiple Choice
1. Desulfovibrio uses sulfate (SO₄²⁻) as its terminal electron acceptor, producing:
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Correct: B) Hydrogen sulfide (H₂S)2. Which bacteria carry out complete ammonia oxidation (comammox) in a single organism?
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Correct: C) Nitrospira inopinata💬 Open-Ended Questions
1. Explain the TCA cycle's dual role as a catabolic and anabolic hub in microbial metabolism. Name at least four biosynthetic precursors generated by the TCA cycle.
Hint / Guidance
Catabolic role: acetyl-CoA (from pyruvate/fatty acids) enters cycle; 2 CO₂ released per turn; 3 NADH + 1 FADH₂ + 1 GTP per turn → energy for ETC. Anabolic (amphibolic) role: intermediates withdrawn for biosynthesis. Biosynthetic precursors: (1) α-ketoglutarate → glutamate family amino acids; (2) Oxaloacetate → aspartate family amino acids, purines; (3) Succinyl-CoA → porphyrins (chlorophyll, haem); (4) Citrate → lipid synthesis (via acetyl-CoA export). Withdrawal of intermediates replenished by anaplerotic reactions (e.g., CO₂ fixation by PEP carboxylase).6. Chemiosmosis
Chemiosmosis — proposed by Peter Mitchell in 1961 — is the mechanism linking electron transport to ATP synthesis. As electrons flow down the ETC, protons are pumped across the membrane, generating a proton-motive force (PMF = Δψ + ΔpH). This PMF drives the rotation of ATP synthase (F₀F₁-ATPase), synthesising ATP from ADP + Pᵢ. Roughly 26 of the ~30 ATP produced per glucose come from chemiosmosis — making the membrane the powerhouse of aerobic respiration.
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1. Nitrogen fixation converts atmospheric N₂ to _______, which can be assimilated by cells.
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Ammonia (NH₃)🔘 Multiple Choice
1. During fermentation, the final electron acceptor is:
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Correct: C) An organic molecule derived from the substrate (e.g., pyruvate → acetaldehyde)2. Nitrifying bacteria such as Nitrosomonas europaea gain energy by oxidising:
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Correct: B) Ammonium (NH₄⁺) to nitrite (NO₂⁻)3. Sulfate-reducing bacteria use sulfate as:
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Correct: C) Terminal electron acceptor in anaerobic respiration💬 Open-Ended Questions
1. What is the significance of alternative electron acceptors in anaerobic respiration for environmental microbiology? Discuss at least three different electron acceptors and their ecological consequences.
Hint / Guidance
(1) NO₃⁻ (denitrification): Pseudomonas etc. reduce NO₃⁻ → N₂O → N₂; removes fixed nitrogen from ecosystems (eutrophication treatment, but reduces soil fertility). (2) SO₄²⁻ (sulfate reduction): Desulfovibrio produces H₂S; drives sulfur cycle, causes anoxic zone odour problems, biocorrosion. (3) Fe³⁺ (iron reduction): Geobacter, Shewanella reduce insoluble Fe(OH)₃ → soluble Fe²⁺; mobilises trace metals and arsenic (groundwater contamination); also bioremediation applications. (4) CO₂ (methanogenesis): Archaea reduce CO₂ → CH₄; key in carbon cycling, greenhouse gas production, wastewater treatment (biogas).7. Protein Catabolism
Protein catabolism provides both energy and biosynthetic precursors during nitrogen or carbon starvation. Extracellular proteases hydrolyse proteins to amino acids; intracellular deamination removes the amino group (→ NH₄⁺, entering the nitrogen cycle), and the remaining carbon skeleton is channelled into central metabolism as pyruvate, acetyl-CoA, oxaloacetate, α-ketoglutarate, or fumarate, depending on the amino acid.
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1. The process by which reduced inorganic compounds such as H₂S are oxidised to provide energy for carbon fixation is called _______.
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Chemolithotrophy🔘 Multiple Choice
1. Chemolithotrophs obtain energy by:
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Correct: C) Oxidising inorganic compounds such as H₂S, NH₃, or Fe²⁺2. Which is the net energy gain difference between aerobic and lactic acid fermentation per glucose, and what accounts for this?
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Correct: A) ~28–30 ATP more from aerobic; accounts for complete oxidation of pyruvate via TCA + ETC + ATP synthase💬 Open-Ended Questions
1. Describe how E. coli adjusts its metabolism when transitioning from aerobic to anaerobic growth. Include changes in gene expression, metabolic pathways used, and the impact on growth rate.
Hint / Guidance
Transition regulated by FNR (Fumarate and Nitrate Reductase) transcription factor — activated under anaerobic conditions. Aerobic → anaerobic: (1) ETC complexes repressed; (2) Anaerobic terminal reductases (nitrate reductase, fumarate reductase) induced; (3) Fermentative pathways activated (mixed acid fermentation — formate, acetate, ethanol, succinate); (4) TCA cycle operates in truncated form; (5) Pyruvate formate lyase replaces pyruvate dehydrogenase. Impact: ATP yield drops from ~30 to 2–17 ATP/glucose; growth rate ~50% slower; biomass yield lower; lactate/formate/ethanol accumulate in medium.2. Trace the path of carbon from CO₂ fixation in the Calvin cycle to the synthesis of glucose. Identify the key enzymes and the energy inputs required.