Chapter 7 Microbial Metabolism
Metabolism (from Greek "metabolan" meaning change) refers to all chemical reactions and physical workings of a cell. It encompasses thousands of reactions, broadly categorized into two main types:
7.1 Metabolism and the Role of Enzymes
Anabolism (Biosynthesis)
Any process that synthesizes cell molecules and structures.
A building and bond-making process.
Forms larger macromolecules from smaller ones.
Usually requires an input of energy (ATP utilized to form bonds).
Examples: Polypeptide synthesis, cell assembly.
Catabolism
The opposite of anabolism.
Breaks the bonds of larger molecules into smaller molecules.
Often releases energy (yields energy).
Degrades macromolecules.
Linking anabolism to catabolism ensures the efficient completion of many cellular processes.
The Critical Role of Electrons in Metabolism
Cells collect and spend energy by transferring electrons from an external source to internal carriers.
These carriers shuttle electrons into a series of proteins that make energy usable for the cell.
Electron flow is key to metabolism.
Metabolism's Three Key Functions
Assembles smaller molecules into larger molecules needed for the cell (Anabolism), utilizing ATP energy.
Degrades macromolecules into smaller molecules (Catabolism), yielding energy.
Collects and spends energy in the form of ATP (Adenosine Triphosphate).
Enzymes: Catalyzing Chemical Reactions
The chemical reactions of life cannot proceed without a special class of molecules called enzymes.
What are Enzymes?
Catalysts: Chemicals that increase the rate of a chemical reaction.
They do not become part of the products or get consumed in the reaction.
Enzymes do not create reactions; they only speed them up.
Characteristics of Enzymes (Table 7.1 Summary)
Made of protein or RNA.
May require cofactors.
Act as organic catalysts to speed up cellular reactions.
Have unique characteristic shape, specificity, and function.
Enable metabolic reactions to proceed at a speed compatible with life.
Have an active site for target substrates.
Are much larger than their substrates.
Associate closely with substrates but do not become integrated into the reaction's products.
Are not used up or permanently changed by the reaction.
Can be recycled, thus functioning in extremely low concentrations.
Greatly affected by temperature and pH.
Can be regulated by feedback loops and genetic mechanisms.
How Enzymes Work
Enzymes speed up metabolic reactions by lowering the activation energy required to initiate the reaction.
Substrates: Molecules acted upon by enzymes.
Enzymes provide a physical site (active site) where substrates can be positioned.
The enzyme is usually much larger than its substrate.
The active site is unique and matches only a particular substrate (Lock and Key model).
An enzyme binds to the substrate, participates directly in changes to the substrate, but does not become part of the products.
It is not used up and can function repeatedly.
Enzyme speeds are very rapid (e.g., lactate dehydrogenase: 8,000 substrates/sec; catalase: 6 million/sec).
Enzyme Structure
Most enzymes are proteins; a special class is made of RNA.
Simple Enzymes: Made of protein alone.
Conjugated Enzymes (Holoenzymes): Contain both protein and non-protein molecules.
Apoenzyme: The actual protein portion.
Cofactors: Non-protein components.
Organic molecules: Called Coenzymes.
Inorganic elements: Metal ions (e.g., iron for catalase).
Enzyme-Substrate Interactions
For a reaction to occur, the substrate must fit closely into the active site. This specific fit is often described as a "lock and key."
The bonds formed between the substrate and enzyme are weak and easily reversible.
Once the enzyme-substrate complex forms, the designated reactions occur on the substrate (often with the aid of a cofactor).
A product is formed and released.
The enzyme can then accept another substrate molecule and repeat the action.
Enzymes can potentially catalyze reactions in both directions, though often depicted in one direction.
Cofactors Supporting Enzyme Work
Many microorganisms require trace elements (specific metal ions) and organic growth factors because these substances act as cofactors for enzymes.
Metallic Cofactors:
Examples: Iron, copper, magnesium, manganese, cobalt, zinc, selenium.
Functions: Activate enzymes, help bring the active site and substrate close together, participate directly in chemical reactions within the enzyme-substrate complex.
Coenzymes:
Organic compounds that work in conjunction with an apoenzyme.
Function: Remove a chemical group from one substrate molecule and add it to another, serving as a transient carrier.
Carry and transfer hydrogen, electrons, carbon dioxide, amino groups, etc.
Many coenzymes are derived from vitamins, explaining their importance in nutrition.
Vitamin deficiencies can prevent holoenzyme formation, compromising chemical reactions and dependent structures/functions.
Naming Enzymes
Classified and named based on site of action, type of action, and substrate.
General Structure: Prefix/stem word (from substrate or reaction type) + suffix "-ase".
Examples:
Carbohydrase: Digests carbohydrate substrates.
Maltase: Digests maltose.
Protease/Peptidase: Breaks peptide bonds of proteins.
Lipase: Digests fats/lipids.
Deoxyribonuclease (DNase): Breaks down DNA.
Synthetase/Polymerase: Bonds small molecules into large ones.
Regulation of Enzyme Action
Enzymes are not all produced in equal amounts or at equal rates.
Constitutive Enzymes
Always present in relatively constant amounts.
Production is not dependent on substrate concentration.
Examples: Enzymes involved in glucose utilization (essential for metabolism).
Regulated Enzymes
Production is turned on (induced) or off (repressed) in response to changes in substrate concentration.
Environmental Factors Affecting Enzyme Activity
Enzymes operate optimally under specific temperature, pH, and osmotic pressure conditions typical of the organism's habitat.
When subjected to changes, enzymes can become chemically unstable or labile.
Low temperatures: Inhibit enzyme reactions.
High temperatures: Denature the apoenzyme.
Denaturation:
Process by which weak bonds maintaining the native shape of the apoenzyme are broken.
Disrupts the enzyme's shape, preventing substrate attachment to the active site.
Leads to non-functional enzymes and failure of metabolic reactions.
Lower/High pH: Can also cause denaturation.
Certain chemicals: Heavy metals, alcohol can act as denaturing agents.
Direct Control of Enzyme Actions
Bacterial cells can directly influence enzyme activity.
Competitive Inhibition
A molecule (mimic) resembles the enzyme's normal substrate.
The mimic occupies the enzyme's active site, preventing the actual substrate from binding.
Since the mimic cannot be acted upon by the enzyme, the enzyme is effectively shut down.
The mimic "competes" with the substrate for the binding site.
Non-Competitive Inhibition (Allosteric Inhibition)
Occurs with enzymes that have two binding sites: an active site and a regulatory site (allosteric site).
A regulatory molecule binds to the regulatory site (not the active site).
This binding changes the conformation (shape) of the active site, preventing the substrate from binding or altering its efficiency.
Often, the regulatory molecule is the product of the enzymatic reaction itself, providing a negative feedback mechanism to slow down activity once a certain product concentration is reached.
Control of Enzyme Synthesis
Controlling enzyme synthesis is another effective mechanism because enzymes do not last indefinitely.
Enzymes are constantly replaced (some are degraded, others diluted by cell division).
Replacement is regulated according to cell demand, involving genetic mechanisms (DNA regulation).
Enzyme Repression
A mechanism to stop further synthesis of an enzyme along its pathway.
As the level of an end product builds up, the genes responsible for producing the enzymes in that pathway are automatically suppressed.
The end product reacts with a site on DNA that regulates enzyme synthesis, inhibiting further production.
Enzyme Induction
The inverse of enzyme repression.
Enzymes appear (are produced) only when suitable substrates are present.
The synthesis of the enzyme is "induced" by its substrate.
Example: E. coli producing lactase only when lactose is present as a carbon source. If sucrose is then provided, lactase synthesis stops, and sucrase synthesis begins.
This enables organisms to utilize various nutrients and prevents wasting energy on enzymes for unavailable substrates.
Metabolic Pathways
Metabolic reactions rarely consist of a single step; they usually occur in multi-step series or pathways.
Each step is catalyzed by a separate enzyme.
The product of one reaction often becomes the reactant (substrate) for the next, forming linear chains.
Many pathways have branches, providing alternate methods for nutrient processing.
Other pathways are cyclic, where the starting molecule is regenerated to initiate another turn.
Pathways are interconnected and merged with others, forming a complex network.
Biochemical Pathway: A specific sequence of reactions where a substrate is converted into a product by the first enzyme, and that product becomes the substrate for the next enzyme, continuing until the final product is made.
Patterns of Metabolism
Metabolic pathways consist of individual chemical reactions that produce intermediate metabolites and lead to a final product.
Patterns include: linear, cyclic, and branched.
Anabolic pathways: Biosynthesis, result in more complex molecules, each step adding a functional group.
Catabolic pathways: Dismantling of molecules, generating energy.
Virtually every reaction in a series involves a specific enzyme.
7.2 Finding and Making Use of Energy
Cells require a constant input of usable energy for metabolic processes.
Energy sources: Sunlight (photosynthesizers), free electrons (electricity-harvesting bacteria), organic substances like sugars (most bacteria).
Energy is primarily stored in ATP.
Energy and Cells
Cells manage energy through chemical reactions that change molecules, often involving bond formation/breaking and electron transfer.
Exergonic Reactions: Release energy as they go forward (energy is on the product side). This energy is available for cellular work.
Example: X + Y + enzyme → C + enzyme + energy
Endergonic Reactions: Require an input of energy to proceed (energy is on the reactant side).
Example: Energy + A + B + enzyme → R + S + enzyme
Exergonic and endergonic reactions are often coupled, so released energy is immediately used.
Cells do not create energy; they extract chemical energy already present in nutrient fuels and apply it to useful work.
Specialized enzyme systems trap energy from nutrient bonds during exergonic reactions.
Energy released is stored in high-energy phosphate bonds, primarily in ATP.
Redox Reactions: Oxidation and Reduction
Some atoms easily give or receive electrons, participating in oxidation-reduction reactions.
Oxidation: The loss of electrons.
Reduction: The gain of electrons.
The compound that loses electrons is oxidized.
The compound that gains electrons is reduced.
Redox Reactions: Oxidation and reduction always occur together in pairs (one substance is oxidized, another is reduced).
Indispensable for cellular energy transformations.
Oxidoreductases: Enzymes that remove electrons from one substrate and add them to another.
Electron Carriers
Oxidoreductases carry coenzymes vital for electron transfer.
Key coenzymes:
NAD+ (Nicotinamide Adenine Dinucleotide)
FAD (Flavin Adenine Dinucleotide)
These coenzymes sit in grooves on enzymes and donate/accept electrons.
Mnemonic: OIL RIG (Oxidation Is Losing, Reduction Is Gaining).
In cellular respiration, oxidation is often the removal of hydrogens (proton + electron); reduction is the addition of hydrogens.
NAD+ as an energy shuttle: During oxidation, an enzyme transfers hydrogen (proton + electron) from a substrate to NAD+, reducing it to NADH. The substrate is oxidized. NADH is a high-energy electron carrier that can donate its hydrogen to other molecules for energy production.
Dehydrogenation: The removal of hydrogens from a compound during a redox reaction.
NAD and FAD function as short-term repositories for electrons until they can be transferred.
They resemble shuttles, alternately accepting and releasing electrons and hydrogens to facilitate redox energy transfer in catabolic pathways.
Electrons are extracted and carried through a series of redox reactions until a final electron acceptor is reached.
Aerobic metabolism: Final acceptor is molecular oxygen.
Anaerobic metabolism: Final acceptor is some other non-oxygen compound.
Adenosine Triphosphate (ATP): Metabolic Money
ATP is the "metabolic money" because it can be earned, saved, spent, and exchanged as energy.
Provides a connection between energy-yielding catabolism and energy-requiring cellular activities.
Structure of ATP
A three-part molecule:
Nitrogenous base (adenine).
Five-carbon sugar (ribose).
Chain of three phosphate groups bonded to the ribose.
The high energy of ATP comes from the orientation of the phosphate groups. They are bulky, negatively charged, and close to each other, creating strain between them (especially the last two).
Removal of the terminal phosphates releases free energy.
Removing the leftmost phosphate yields ADP (Adenosine Diphosphate).
Removing the next phosphate yields AMP (Adenosine Monophosphate).
AMP derivatives also form the backbone of RNA and are components of coenzymes like NAD and FAD.
Metabolic Role of ATP
Primary energy currency of the cell.
Used and replenished in an ongoing cycle.
Energy released during ATP hydrolysis often drives biosynthesis by providing an activating phosphate to a substrate (phosphorylation).
ATP is also used to prepare molecules for catabolism (e.g., phosphorylation of glucose in glycolysis).
When the terminal phosphate is used, ATP becomes ADP + energy.
To replenish ATP, a terminal phosphate group is added back to ADP, requiring an input of energy.
In heterotrophs, this energy comes from catabolic pathways where nutrients (e.g., carbohydrates) are degraded.
ATP is formed when substrates or electron carriers provide a high-energy phosphate that bonds to ADP.
7.3 Catabolism
Catabolism uses enzymes to break down organic molecules into precursor molecules that cells can then use to build larger, more complex molecules (anabolism).
Reducing Power: Electrons are available from NADH and FADH2 (reduced forms).
Energy Storage: Energy is stored in ATP bonds.
Both reducing power and ATP are needed in large quantities for anabolic parts of metabolism.
Catabolism starts with nutrients from the environment, which cells acquire via transfer mechanisms (some requiring energy from existing catabolism).
Intracellular nutrients are then broken down into appropriate precursor molecules via catabolic pathways.
Nutrient Processing Pathways
Nutrient processing is varied, especially in bacteria, but often centers on three catabolic pathways, frequently using glucose as the nutrient.
Overview of Catabolic Pathways
Aerobic Respiration:
Series of reactions (glycolysis, Krebs cycle, respiratory chain) that convert glucose to CO2.
Recovers significant amounts of energy (large ATP yield).
Relies on free oxygen as the final electron acceptor.
Characteristic of many bacteria, fungi, protozoa, and animals.
Anaerobic Respiration:
Uses the same three pathways as aerobic respiration (glycolysis, Krebs cycle, ETS).
Does not use oxygen as the terminal electron acceptor.
Instead, uses inorganic compounds like NO3-, SO42-, CO32-, etc.
Variable ATP yield (less than aerobic respiration).
Fermentation:
An adaptation for facultative and aerotolerant anaerobes.
Incompletely oxidizes glucose.
Oxygen is not required.
Organic compounds are the terminal electron acceptors.
Produces relatively small amounts of ATP.
Creates useful byproducts.
Glycolysis
All three metabolic pathways begin with glycolysis.
Converts glucose into two molecules of pyruvic acid.
Pyruvic acid is a chemical uniquely capable of yielding energy in subsequent pathways.
Process:
Initial steps: Two ATP molecules are used to add two phosphates to glucose, forming a 6-carbon sugar diphosphate.
This 6-carbon molecule splits into two 3-carbon molecules.
Each 3-carbon molecule is converted to pyruvate through a series of steps.
Energy Release: During these reactions, electrons are transferred to 2 NAD+ to form 2 NADH, and 4 ATP molecules are formed.
Net Yield (from one glucose molecule):
2 molecules of Pyruvate
2 NADH
2 ATP (4 made, 2 used)
Under aerobic conditions, pyruvate is further oxidized for more ATP.
Under anaerobic conditions, pyruvate is converted into other organic products (acids or alcohols).
Pyruvic Acid: Central to Metabolism
In strictly aerobic organisms and some anaerobes, pyruvic acid enters the Krebs cycle.
Facultative anaerobes can use fermentation, where pyruvic acid is reduced to acids or other products.
Bacterial respiration (aerobic or anaerobic) utilizes the Krebs cycle and the electron transport system.
Fermentation does not use the Krebs cycle or the electron transport system.
The Krebs Cycle (Tricarboxylic Acid Cycle - TCA Cycle)
A carbon and energy wheel.
Glycolysis yields a small amount of energy and pyruvic acid, which is still energy-rich.
The Krebs cycle is the second phase of catabolism, occurring in the cytoplasm of bacteria and the mitochondrial matrix in eukaryotes.
Process (for one pyruvate molecule):
Pyruvate (3C) is converted to Acetyl-CoA (2C).
CO2 is produced.
1 NADH is formed.
Acetyl-CoA (2C) enters the Krebs cycle by combining with oxaloacetate (4C) to form citrate (6C). CoA is released.
Citrate undergoes a series of transformations:
Two decarboxylations (release CO2).
Four oxidations (transfer H and electrons to NAD+ and FAD).
This regenerates oxaloacetate (4C), making it a cycle.
Yield (from one Acetyl-CoA, i.e., half of one glucose molecule):
2 CO2
3 NADH
1 FADH2
1 ATP
Total Yield (from one glucose molecule, requiring two turns of the cycle):
4 CO2
6 NADH
2 FADH2
2 ATP
The main products are reduced coenzymes (NADH and FADH2), which are vital for subsequent energy production in the electron transport chain.
The Respiratory Chain (Electron Transport System - ETS)
The final processing mill for electrons and hydrogen ions, and the major generator of ATP.
Final step in both aerobic and anaerobic respiration.
Consists of a chain of special redox carriers.
Receives electrons from reduced carriers (NADH and FADH2) generated by glycolysis and the Krebs cycle.
Passes electrons sequentially and orderly from one redox molecule to the next.
Components:
NADH dehydrogenase
Flavoproteins
Coenzyme Q (Ubiquinone)
Cytochromes (contain a tightly bound metal atom that accepts/donates electrons).
In bacteria, ETS carriers and enzymes are embedded in the cytoplasmic membrane.
In eukaryotes, they are in the inner mitochondrial membrane.
Mechanism:
Electrons from NADH and FADH2 are transferred to carriers at the beginning of the chain.
As electrons move down the chain (in a series of redox reactions), protons (H+) are pumped out of the membrane, creating a concentration gradient (more protons outside than inside).
This creates a proton motive force (PMF). The extracellular space becomes more positively charged and acidic.
Protons flow back into the cell, down their concentration gradient, through specialized protein channels called ATP Synthase.
The energy from this proton flow is used to synthesize ATP from ADP and phosphate. This process is called Oxidative Phosphorylation.
At the end of the chain, the electrons and protons are accepted by a terminal electron acceptor.
Terminal Electron Acceptor:
Aerobic Respiration: Molecular oxygen (O2). Oxygen accepts electrons and hydrogen to produce water (H2O). This consumes oxygen.
Anaerobic Respiration: Non-oxygen compounds (e.g., nitrate, sulfate).
ATP Yield:
Each NADH entering the ETS can yield a maximum of 3 ATPs (actual numbers may be lower due to inefficiencies).
Each FADH2 entering the ETS (at a later point) can yield a maximum of 2 ATPs.
Aerobic Respiration: Maximum of 38 ATPs per glucose.
Anaerobic Respiration: Yields less ATP than aerobic respiration (2 to 36 ATPs).
Toxic Oxygen Products:
Incomplete reduction of oxygen can produce toxic byproducts like superoxide ion (O2-) and hydrogen peroxide (H2O2).
Aerobes have neutralizing enzymes: superoxide dismutase and catalase.
Genus Streptococcus lacks cytochromes and catalase but can grow in oxygen due to a special peroxidase.
Anaerobes generally lack these enzymes, limiting their ability to process free oxygen and contributing to its toxic effects on them.
Anaerobic Respiration
Terminal step utilizes inorganic compounds (not O2) as the final electron acceptor in the ETS.
Best-known systems: Nitrate (NO3-) and nitrite (NO2-) reduction.
Example: E. coli uses nitrate reductase to remove oxygen from nitrate, producing nitrite and water.
Denitrification: Some bacteria (e.g., Pseudomonas, Bacillus) can further reduce nitrate to nitric oxide, nitrous oxide, and even nitrogen gas, important for nitrogen recycling.
Other inorganic acceptors: Carbonates, sulfates.
Facultative anaerobes can switch between aerobic and anaerobic respiration based on oxygen availability. Aerobic respiration is more efficient.
Fermentation
Definition: Incomplete oxidation of glucose or other carbohydrates in the absence of oxygen.
Uses organic compounds as terminal electron acceptors.
Yields a small amount of ATP (maximum 2 ATP per glucose).
Used by organisms that lack an electron transport chain or when oxygen is absent.
Allows independence from molecular oxygen, enabling colonization of anaerobic environments.
Allows microbes to adapt to variations in oxygen availability.
Rapid growth can be maintained by increasing the rate of glycolysis.
Examples:
Human muscle cells: Convert pyruvic acid to lactic acid during strenuous activity when oxygen is depleted, allowing short-term anaerobic ATP production (causes muscle fatigue).
Cattle rumen bacteria: Ferment cellulose to organic acids.
Products of Fermentation:
Pyruvic acid can become an electron acceptor and proceed through:
Alcoholic fermentation: Produces ethanol and CO2 (e.g., in beverages, bread dough).
Acidic fermentation: Produces various organic acids (e.g., lactic acid, acetic acid, propionic acid, butanol).
Organic molecules reduced in fermentation are varied and yield useful products.
Microbes are harnessed to synthesize vitamins, antibiotics, hormones (e.g., hydrocortisone).
Large-scale industrial processes are often called "fermentation" even if they occur aerobically.
Catabolism of Non-Carbohydrate Compounds
Besides glucose, other compounds serve as fuel.
Complex Polysaccharides: Broken down into component sugars, which enter glycolysis at various points.
Lipids (Fats):
Broken down by lipases into fatty acids and glycerol.
Glycerol is converted to dihydroxyacetone phosphate (DHAP), which enters glycolysis.
Fatty acids undergo beta-oxidation: 2-carbon units are successively transferred to Coenzyme A, creating Acetyl-CoA, which enters the Krebs cycle.
Beta-oxidation yields a large amount of energy (e.g., 6-carbon fatty acid yields 50 ATPs vs. 38 for a 6-carbon sugar).
Proteins:
Broken down by proteases into amino acid components.
Amino groups are removed by deamination.
The remaining carbon compound is converted to one of several Krebs cycle intermediates.
7.4 Anabolism and the Crossing Pathways of Metabolism
This section focuses on anabolic functions, biosynthesis, and the interconnectedness of metabolic pathways.
Amphibolism
Cells carefully manage carbon compounds.
Most catabolic pathways contain strategic molecular intermediates that can be diverted into anabolic pathways.
This allows a given molecule to serve multiple purposes, maximizing benefit from nutrients.
Amphibolism: The ability of a system to integrate catabolic and anabolic pathways to improve cell efficiency.
Examples of amphibolic pathways (e.g., glucose metabolism):
Intermediates from glycolysis can be fed into amino acid synthesis, then protein formation.
Amino acids contribute nitrogenous groups to nucleotides for nucleic acids.
Monosaccharides (like glucose) are made into other sugars and polymerized into complex carbohydrates.
Glycolysis product Acetyl-CoA can be oxidized to form fatty acids (lipids).
Amino acids can be deaminated, leading to intermediates like pyruvate and Acetyl-CoA.
Fatty acids can be oxidized to form Acetyl-CoA.
Anabolism: Formation of Macromolecules
Building blocks (monosaccharides, amino acids, fatty acids, nitrogen bases, vitamins) make up macromolecules and organelles.
Sources of Building Blocks
Enter the cell from the outside (ready to use).
Synthesized through various cellular pathways.
The degree of synthesis capability varies by organism's genetic makeup.
Autotrophs: Require only CO2 as a carbon source and a few minerals to synthesize all cell substances.
Heterotrophs: Some (e.g., E. coli, yeast) are efficient, synthesizing all cellular substances from minerals and a single organic carbon source (like glucose). Others (strict parasites) have few synthetic abilities and derive most precursors from the host.
Once building blocks are in the "metabolic pool," they are available for polymer synthesis.
Carbohydrate Biosynthesis
Glucose has a central role in metabolic energy and is crucial for cell structures (e.g., cellulose in cell walls, storage granules like starch/glycogen).
An intermediate in glycolysis, glucose-6-phosphate, is used to form glycogen.
Other monosaccharides are important for bacterial cell walls (e.g., peptidoglycan contains muramic acid and glucosamine, derived from fructose-6-phosphate).
Deoxyribose and ribose are essential building blocks for nucleic acids.
Polysaccharides are predominant components of cell surface structures (capsules, glycocalyx, slime layers).
Amino Acid and Protein Synthesis
Proteins account for a large portion of cell constituents (enzymes, membranes, cell walls, appendages).
Generally, 20 amino acids are needed for proteins.
Some organisms (e.g., E. coli) can synthesize all 20.
Others (e.g., animals) lack some or all pathways and must acquire essential amino acids from their diet.
Protein synthesis is a complex process requiring genetic blueprints and intricate cellular machinery (covered in Chapter 8).
Medical Moment: Essential, Non-Essential, and Conditionally Essential Amino Acids
Essential Amino Acids: Must be obtained from the diet; bodies cannot synthesize them.
Non-Essential Amino Acids: Can be synthesized by the body.
Conditionally Essential Amino Acids: Not normally required in the diet, but become essential in specific populations unable to synthesize them adequately.
Example: Individuals with Phenylketon