Microbial Metabolism: Structured Study Guide

I Microbial Metabolism

What Is Metabolism?

Metabolism encompasses all chemical reactions occurring within a cell, enabling life processes. These reactions are organized into distinct pathways:

• Catabolic pathways: Break down molecules and release energy.

• Anabolic pathways: Build molecules and require energy; also known as biosynthetic reactions.

• Amphibolic pathways: Can function in both catabolism and anabolism.

Key connection: Catabolism releases energy, which is used to generate ATP. ATP then powers anabolic reactions.

ATP — The Cell's Energy Molecule

ATP (adenosine triphosphate) is the primary energy carrier in cells, consisting of adenine, ribose, and three phosphate groups. ADP (adenosine diphosphate) contains two phosphate groups.

• Dephosphorylation: Removal of the terminal phosphate from ATP releases energy and produces ADP.

• Phosphorylation: Addition of a phosphate to ADP recharges it to ATP.

• Cells continuously recycle ATP and ADP rather than storing large amounts of ATP.

Equation: $\mathrm{ATP} \xrightarrow{\text{dephosphorylation}} \mathrm{ADP} + \text{energy}$ $\mathrm{ADP} + \text{phosphate} \xrightarrow{\text{phosphorylation}} \mathrm{ATP}$

Enzymes

Enzymes are protein catalysts that facilitate cellular reactions by lowering activation energy. They are not consumed during the reactions they catalyze.

• The substrate binds at the enzyme's active site, forming an enzyme-substrate complex.

• After the reaction, the enzyme is available for reuse.

• Enzyme activity is sensitive to temperature and pH; extreme conditions can denature enzymes.

• Cofactor: Non-protein helper required by some enzymes.

• Coenzyme: Organic cofactor (e.g., NAD+ and FAD).

• Ribozyme: Catalytic RNA molecule.

Enzyme regulation:

• Feedback inhibition: End product reduces pathway activity.

• Competitive inhibition: Inhibitor competes for the active site.

• Noncompetitive inhibition: Inhibitor binds elsewhere, altering enzyme activity.

• Allosteric regulation: Molecule binds at a regulatory site, changing enzyme activity.

Redox Reactions

Redox reactions are fundamental to energy harvesting in cells. Oxidation is the loss of electrons, while reduction is the gain of electrons. These reactions are always coupled.

• Memory trick: OIL RIG — Oxidation Is Loss, Reduction Is Gain.

• Example: $\mathrm{Fe} + \mathrm{Cu}^{2+} \rightarrow \mathrm{Fe}^{2+} + \mathrm{Cu}$

• Cells harvest energy by oxidizing nutrients; sugars and fats are highly reduced and release energy when oxidized.

NAD+ and NADH

NAD+ is the oxidized form, while NADH is the reduced form. These molecules act as electron carriers in metabolic pathways.

• NAD+ is available to accept electrons.

• NADH is carrying electrons.

Three Ways Cells Recharge ADP to ATP

Cells use three mechanisms to regenerate ATP from ADP:

Mechanism	How ATP is made	ETC?	Examples
Substrate-level phosphorylation	High-energy phosphate transferred directly to ADP	No	Glycolysis, Krebs cycle, some fermentation
Oxidative phosphorylation	ADP recharged using electron transport chains powered by nutrients	Yes	Aerobic and anaerobic respiration
Photophosphorylation	ADP recharged using electron transport chains powered by solar energy	Yes	Light-dependent reactions of photosynthesis

Cellular Respiration

Cellular respiration is a multi-step process involving glycolysis, the intermediate step, the Krebs cycle, and the electron transport chain (ETC).

Glycolysis

• Occurs in the cytoplasm.

• Does not require oxygen.

• Consists of 10 reactions: energy investment and payoff stages.

• Glucose is split into two molecules of pyruvic acid.

• ATP is produced via substrate-level phosphorylation.

Intermediate Step

• Each pyruvic acid is converted to acetyl-CoA.

• Occurs in cytoplasm (prokaryotes) or mitochondrial matrix (eukaryotes).

• Transitory step between glycolysis and Krebs cycle.

Krebs Cycle

• Also called citric acid cycle or tricarboxylic acid cycle.

• Includes redox and decarboxylation reactions.

• Produces ATP and reduced cofactors (NADH, FADH2).

• Cycle runs twice per glucose molecule.

• Carbons from glucose are released as CO2.

Electron Transport Chain (ETC)

• NADH and FADH2 donate electrons to the ETC.

• Electron movement supports formation of a proton gradient.

• Proton gradient is used to make ATP.

Where Cellular Respiration Occurs

Process	Prokaryotic cell	Eukaryotic cell
Glycolysis	Cytoplasm	Cytoplasm
Intermediate step	Cytoplasm	Mitochondrial matrix
Krebs cycle	Cytoplasm	Mitochondrial matrix
Electron transport chain	Plasma membrane	Inner mitochondrial membrane

Fermentation

Fermentation is an anaerobic pathway that works with glycolysis and does not use a respiratory chain. Its main purpose is to transfer electrons from NADH to an organic molecule and regenerate NAD+ so glycolysis can continue.

• Occurs in the cytoplasm.

• Does not use an electron transport chain.

• ATP production is mainly associated with glycolysis.

Fermentation Pathways

Pathway	Products / Key Point
Homolactic	Lactic acid
Alcohol	Ethanol
Mixed acid	Ethanol, gases, acids (succinic, acetic, formic), CO2
Butanediol	Acetoin and butanediol; VP test detects acetoin

Fermentation vs. Respiration

Fermentation	Cellular Respiration
No respiratory chain	Uses a respiratory chain
Occurs in cytoplasm	Uses multiple cellular locations
Regenerates NAD+ so glycolysis can continue	Uses electron transport to transfer electrons
Anaerobic	Can be aerobic or anaerobic

Additional info: Anaerobic respiration uses an electron transport chain, unlike fermentation.

Using Metabolic Properties to Identify Bacteria

Bacterial species possess unique metabolic profiles, which serve as biochemical fingerprints for identification. Biochemical tests detect metabolic end products, intermediates, or specific enzymes.

• Over 1,000 tests are available for microbial identification.

• Tests range from specialized media to molecular, genetic, and metabolic procedures.

Identifying an Unknown Bacterial Specimen

1. Prepare a pure culture.

2. Use staining, microscopy, and general culture characteristics to narrow possibilities.

3. Subject the culture to a panel of biochemical tests.

Important Biochemical Tests

• Amino acid catabolism: Tests for deaminases and decarboxylases (e.g., phenylalanine deaminase, ornithine decarboxylase).

• Sulfur reduction / H2S: Catabolism of sulfur-containing amino acids releases H2S, which forms a black precipitate with iron.

• Fermentation tests: Media contains protein, carbohydrate, pH indicator, and sometimes a Durham tube. Acid production lowers pH and changes indicator color; gas is captured in Durham tube.

• MR / VP: MR tests for mixed-acid fermentation; VP detects acetoin (butanediol fermentation intermediate).

• Oxidase and catalase: Oxidase test detects cytochrome c oxidase (purple/blue pigment); catalase test detects catalase (bubbles from oxygen release).

• API system: Analytical Profile Index; positive results generate an index number matched to bacterial candidates in a database.

Test-Day Quick Review

• Metabolism: All chemical reactions in a cell

• Catabolism: Breaks down molecules; releases energy

• Anabolism: Builds molecules; uses energy

• ATP: Cellular energy molecule

• Dephosphorylation: Removes phosphate; releases energy

• Phosphorylation: Adds phosphate; recharges ATP

• Enzyme: Protein catalyst

• Oxidation: Loses electrons

• Reduction: Gains electrons

• OIL RIG: Oxidation Is Loss; Reduction Is Gain

• NAD+: Oxidized electron carrier

• NADH: Reduced electron carrier

• Glycolysis: Cytoplasm; does not require oxygen

• Intermediate step: Pyruvic acid → acetyl-CoA + CO2

• Krebs cycle: Runs twice per glucose

• Eukaryotic ETC: Inner mitochondrial membrane

• Prokaryotic ETC: Plasma membrane

• Fermentation: No ETC; regenerates NAD+

• Homolactic: Lactic acid

• Alcohol fermentation: Ethanol

• VP: Detects acetoin

• MR: Tests mixed-acid fermentation

• H2S: Black precipitate with iron

• Catalase positive: Bubbles

• Oxidase: Tests cytochrome c oxidase

• API: Uses an index number to aid bacterial identification

One Big Picture

Nutrients → Catabolism → Redox reactions → NAD+/NADH → ATP production → ATP powers cellular work and anabolism. Glucose → Glycolysis → Pyruvic acid → Intermediate step → Acetyl-CoA → Krebs cycle → NADH/FADH2 → ETC → ATP If ETC cannot operate: Glucose → Glycolysis → NADH → Fermentation → NAD+ regenerated → Glycolysis continues.

Study priority: Know the differences between catabolism/anabolism, oxidation/reduction, phosphorylation mechanisms, the locations of respiration steps, fermentation vs. respiration, and what the bacterial identification tests detect.