Microbial Metabolism, Growth, and Genetics: Study Guide (Chapters 5-7)

Microbial Metabolism

Catabolism vs. Anabolism

• Catabolism: The breakdown of larger molecules into smaller products. These reactions are exergonic, meaning they release energy (often captured as ATP).

• Anabolism: The synthesis of large molecules from smaller ones. These reactions are endergonic, requiring energy input, typically from ATP.

• Metabolism: The sum of all chemical reactions in a cell, including both catabolic and anabolic pathways. Example: Breakdown of glucose and synthesis of peptidoglycan for cell reproduction.

• Amphibolic reactions: Reactions that can proceed in both catabolic and anabolic directions, providing metabolic flexibility.

Key Definitions

• Exergonic: Reactions that release energy.

• Endergonic: Reactions that require energy input.

ATP: Structure and Function

• Adenosine triphosphate (ATP) is the primary energy currency of the cell.

• Structure: Adenine base, ribose sugar, and three phosphate groups.

• Function: Stores and transfers energy for cellular processes.

Enzymes: Structure, Function, and Regulation

• Enzymes are biological catalysts that speed up chemical reactions by lowering activation energy.

• Structure: Most are proteins with an active site for substrate binding.

• Factors affecting enzyme activity: Temperature, pH, substrate concentration, and presence of inhibitors.

• Denaturation: Loss of enzyme structure (and function) due to extreme conditions.

Enzyme Inhibition

• Allosteric inhibition: Inhibitor binds to a site other than the active site, changing enzyme shape.

• Competitive inhibition: Inhibitor competes with substrate for the active site.

• Noncompetitive inhibition: Inhibitor binds elsewhere, reducing enzyme activity regardless of substrate concentration.

• Feedback inhibition: End product of a pathway inhibits an earlier enzyme, regulating pathway activity.

• Sulfanilamide acts as a competitive inhibitor, blocking the enzyme that synthesizes folic acid in bacteria.

Activation Energy

• Activation energy: The minimum energy required to initiate a chemical reaction.

Redox Reactions and Coenzymes

• Oxidation-reduction (redox) reactions: Transfer of electrons between molecules; essential for energy extraction.

• NADH and FADH2: Electron carriers that shuttle electrons to the electron transport chain.

Carbohydrate Catabolism Pathways

• Glycolysis: Occurs in cytoplasm; glucose is split into two pyruvate molecules, producing ATP and NADH.

• Acetyl-CoA synthesis: Pyruvate is converted to acetyl-CoA, releasing CO2 and generating NADH.

• Citric Acid Cycle (Krebs Cycle): Acetyl-CoA is oxidized, producing NADH, FADH2, ATP, and CO2.

• Electron Transport Chain (ETC): Electrons from NADH and FADH2 are transferred through membrane proteins, generating a proton gradient used to produce ATP.

Location of Pathways

• In prokaryotes: All steps occur in the cytoplasm or plasma membrane.

• In eukaryotes: Glycolysis in cytoplasm; acetyl-CoA synthesis and Krebs cycle in mitochondria; ETC in mitochondrial inner membrane.

ATP Production

• Substrate-level phosphorylation: Direct transfer of phosphate to ADP during glycolysis and Krebs cycle.

• Oxidative phosphorylation: ATP synthesis powered by the proton gradient generated by the ETC (chemiosmosis).

Fermentation

• Occurs when oxygen is absent; regenerates NAD+ for glycolysis.

• Types: Alcoholic fermentation (produces ethanol and CO2), lactic acid fermentation (produces lactic acid).

• Purpose: To replenish NAD+ for continued glycolysis.

Catabolism of Fats and Proteins

• Beta-oxidation: Fatty acids are broken down into acetyl-CoA units for entry into the Krebs cycle.

• Deamination: Removal of amino groups from proteins, allowing carbon skeletons to enter central catabolic pathways.

• Carbohydrate catabolism is called Central Catabolism because it provides intermediates for other macromolecule breakdowns.

Integration and Regulation of Metabolism

• Cells regulate metabolism via enzyme activity, gene expression, and feedback inhibition to maintain homeostasis.

Microbial Nutrition and Growth

Oxygen Requirements and Energy Metabolism

• Obligate aerobe: Requires oxygen for growth; uses aerobic respiration.

• Obligate anaerobe: Cannot tolerate oxygen; uses anaerobic respiration or fermentation.

• Facultative anaerobe: Grows with or without oxygen; prefers aerobic respiration but can switch to fermentation or anaerobic respiration.

• Aerotolerant anaerobe: Does not use oxygen but tolerates its presence.

• Microaerophile: Requires low levels of oxygen.

• Tested using fluid thioglycolate broth (FTM) to observe growth patterns.

Protection from Toxic Oxygen

• Microbes use enzymes to detoxify reactive oxygen species:

  • Superoxide dismutase: Converts superoxide radicals to hydrogen peroxide.

  • Catalase: Converts hydrogen peroxide to water and oxygen.

  • Peroxidase: Reduces hydrogen peroxide to water.

Binary Fission

• Primary method of prokaryotic cell division.

• Steps: DNA replication, cell elongation, septum formation, and cell separation.

• Differs from mitosis (eukaryotic division) as it does not involve spindle formation or multiple chromosomes.

• Results in two genetically identical daughter cells.

Nutritional Requirements for Growth

• Essential nutrients: Carbon, nitrogen, sulfur, phosphorus, trace elements, and growth factors.

• Cells use these nutrients for biosynthesis, energy production, and cellular maintenance.

Phases of Microbial Growth

• Lag phase: Cells adapt to environment; no division.

• Log (exponential) phase: Rapid cell division; population doubles at a constant rate.

• Stationary phase: Nutrient depletion and waste accumulation slow growth; cell division equals cell death.

• Death phase: Cells die faster than they divide.

• Relates to lab techniques (e.g., Gram stain) and antibiotic sensitivity (most effective during log phase).

Trophic Classification of Microbes

• Chemoheterotrophs: Obtain energy and carbon from organic compounds.

• Photoautotrophs: Use light for energy and CO2 as carbon source.

• Chemoautotrophs: Use inorganic chemicals for energy and CO2 as carbon source.

• Photoheterotrophs: Use light for energy and organic compounds for carbon.

Environmental Effects on Growth

• Temperature: Human pathogens grow best at 35–37°C (mesophiles).

• pH: Most bacteria prefer neutral pH (6.5–7.5).

• Osmotic pressure: High salt or sugar inhibits growth; halophiles tolerate high salt.

Bacterial Population Growth

• Exponential (logarithmic) growth: Population doubles each generation.

• Arithmetic growth: Population increases by a constant number each generation (rare in microbes).

• Calculation: $N = N_0 \times 2^n$ Where $N$ = final cell number, $N_0$ = initial cell number, $n$ = number of generations.

Toxic Forms of Oxygen and Protective Enzymes

Toxic Oxygen Form	Protective Enzyme
Singlet oxygen	Carotenoids
Superoxide radical (O2-)	Superoxide dismutase
Peroxide anion (O22-)	Catalase, Peroxidase
Hydroxyl radical (OH•)	No enzyme; minimized by other systems

Biofilms

• Complex communities of microbes attached to surfaces and embedded in a self-produced matrix.

• Clinically important due to increased resistance to antibiotics and immune responses.

• Quorum sensing: Cell-to-cell communication that coordinates gene expression and biofilm formation.

Culturing Bacteria

• Defined media: Exact chemical composition known.

• Complex media: Contains extracts; composition varies.

• Differential media: Distinguishes between organisms based on biochemical reactions.

• Selectivemedia: Inhibits growth of some organisms while allowing others.

Pure Culture Techniques

• Streak plate and pour plate methods isolate single colonies.

• Colony-forming unit (cfu): A single cell or group of cells that gives rise to a colony.

• Axenic culture: Pure culture containing only one species.

• Aseptic technique: Procedures to prevent contamination.

Measuring Microbial Growth

• Direct count: Counting cells under a microscope or using a cell counter.

• Viable count: Counting colonies formed on agar plates.

• Turbidity measurement: Using a spectrophotometer to estimate cell density based on light absorption.

Microbial Genetics

Ames Test

• Detects mutagenic potential of chemicals using Salmonella auxotrophs (cannot synthesize histidine).

• Positive result: Reversion to prototrophy indicates mutation.

• Does not indicate carcinogenicity in humans directly.

• Auxotroph: Mutant organism lacking the ability to synthesize a particular nutrient.

Mutations

• Point mutations: Single nucleotide changes.

• Silent mutation: No change in amino acid sequence.

• Missense mutation: Changes one amino acid.

• Nonsense mutation: Introduces a stop codon, truncating the protein.

• Frameshift mutations: Insertions or deletions that shift the reading frame.

Structure of Nucleotides and Nucleic Acids

• Nucleotide: 5-carbon sugar, phosphate group, nitrogenous base.

• DNA: Deoxyribose sugar, double-stranded, bases A, T, C, G.

• RNA: Ribose sugar, single-stranded, bases A, U, C, G.

• Types of RNA: mRNA (messenger), tRNA (transfer), rRNA (ribosomal).

Genetic Code and Central Dogma

• Replication: DNA is copied by DNA polymerase (semiconservative process).

• Transcription: DNA is transcribed to RNA by RNA polymerase.

• Translation: mRNA is decoded to synthesize proteins at the ribosome.

• Enzymes: DNA polymerase, RNA polymerase, ribosomes, tRNA synthetase.

Mutagens

• Mutagen: Agent that increases mutation rate (e.g., chemicals, radiation).

• Results: Can cause beneficial, neutral, or harmful mutations.

Horizontal Gene Transfer in Bacteria

• Transformation: Uptake of naked DNA from the environment.

• Transduction: Transfer of DNA by bacteriophages (viruses); can be generalized or specialized.

• Conjugation: Direct transfer of DNA via cell-to-cell contact, often involving a pilus (F pilus).

Chromosomes and Plasmids

• Prokaryotic chromosomes: Usually single, circular DNA molecule.

• Eukaryotic chromosomes: Multiple, linear DNA molecules.

• Plasmids: Small, circular DNA molecules; replicate independently; may carry antibiotic resistance genes (e.g., F plasmid for conjugation).

Genetic Recombination

• Exchange of genetic material between DNA molecules; increases genetic diversity.

• Occurs naturally during horizontal gene transfer and in eukaryotes during meiosis.

Proofreading Error Rates

• DNA polymerase: High fidelity, low error rate due to proofreading activity.

• RNA polymerase: Higher error rate; lacks proofreading.

Operons: Inducible vs. Repressible

• Operon: Cluster of genes under control of a single promoter.

• Inducible operon: Usually off; activated by substrate (e.g., lac operon).

• Repressible operon: Usually on; repressed by end product (e.g., trp operon).

• Promoter: DNA sequence where RNA polymerase binds to initiate transcription.

Lactose (lac) Operon

• Controls metabolism of lactose in Escherichia coli.

• Structure: Promoter, operator, and three structural genes (lacZ, lacY, lacA).

• Function: Induced in presence of lactose; repressed when lactose is absent.

• Role in homeostasis: Allows bacteria to adapt to nutrient availability.