Microbial Metabolism

Metabolic Reactions Enzymology Catabolism Phototrophy Anabolism

Oxidation;

e

- loss to acceptor

Metabolism Overview:

Reduction;

e

- gain from donor

Metabolic Pathways

• Although we can recognize substrate and product of individual enzymatic reactions; metabolic functions are often performed by several enzymatic reactions in a series or “pathway”.

• Pathways can be linear, branched, cyclic or even spiral.

• Pathway activity is controlled in three ways: – Metabolites and enzymes may be localized in different parts of the cell; called metabolic channeling. (important in eukaryotes) – The total amount of enzymes in a pathway can vary (gene expression).

– Pathway activity is controlled by critical regulated enzymes. These “pacemaker enzymes” are often the rate-limiting step in the pathway.

“Pacemaker” Enzyme Activity

• Enzyme activity may change due to inhibitor or activator molecules called effectors.

• Inhibitors can be competitive (bind at substrate active site) • Noncompetitive inhibitors and activators bind to allosteric (regulatory) sites; separate from the active site; These effectors change the shape of the protein and its activity as a catalyst.

Metabolic Pathway Control Strategies

Feedback Inhibition: (“end-product inhibition”; red) • rate limiting enzyme is first in pathway and is allosteric.

• end-product is a negative effector (inhibitor) of first enzyme Feed Forward Activation: (“earlier-substrate activation”; blue) • rate limiting enzyme of a branch point is allosteric.

• earlier-substrate is a positive effector (activator) of a forward reaction enzyme.

+ Arrows = enzymes

Reversible Metabolic Pathways

• Amphibolic Pathways: – Catabolic direction – Anabolic direction • Separate regulatory enzymes each way function as “check valves” for flow control.

• Other pathway enzymes are reversible; their equilibrium shifts based on concentration of reactants & products.

• Gycolysis / Gluconeogenesis is a good example. Catabolic breakdown of glucose for energy versus the its anabolic formation, respectively.

Glucose Catabolism

• ATP as the cellular energy storage unit, can be formed during respiration (R) or fermentation (F).

• Both contain the Glycolysis pathway; which produces ATP, the electron carrier molecule NADH, and pyruvate from glucose.

• Aerobic Respiration will proceed via Krebs Cycle and an ETC if there is oxygen to react as a terminal electron acceptor.

• Oxygen is not the only possible terminal electron acceptor in some bacteria (e.g. NO 3 or SO 4 can be used); called Anaerobic Respiration.

(ETC) (aerobic) (anaerobic)

• Fermentation proceeds when there is

no

terminal electron acceptor for respiration.

Products of Fermentation

Without any form of respiration, glycolysis products, pyruvate and NADH, will accumulate. To keep making any more ATP by glycolysis, fermenting cells must convert NADH (red.) back to NAD+ (ox.) by passing its electrons to pyruvate. Reaction pathways that do this convert pyruvate to many other compounds, depending on the organism.

Glycolysis:

6C glucose goes to 2x 3C pyruvate plus 2 ATP net, and 2 NADH. ATP must first be invested to then yield energy from oxidation and substrate level phosphorylation of ATP.

Pyruvate Decarboxylation:

(Preparatory Step Before Kreb Cycle) • Pyruvate loses a carbon in the form of CO 2 ; an electron is removed to convert NAD+ to NADH, and coenzyme-A (CoA) binds to the 2C acetyl group.

• Acetyl CoA enters the Kreb Cycle by binding with 4C oxaloacetate to form 6C citric acid.

Krebs Cycle:

• The cycle converts a citric acid back to oxaloacetate; losing 2 CO 2 ; releasing electrons to yield 3 NADH plus 1 FADH, and one ATP by substrate level phosphorylation.

• For one glucose the cycle runs twice.

Energy Perspective on the Electron Transport Chain (ETC) Function

The ETC is a series of membrane bound electron carriers that transports electrons from high to low energy state, ending with oxygen accepting electrons to water.

Energy release is first used to pump protons (H+) across the membrane; a proton motive force (PMF) then drives ATP synthesis.

Each NADH will make 3 ATP. Each FADH will make 2 ATP

Energy State

Each electron transport step releases energy

FADH

PMF= more protons on this side of membrane.

Only 2 ATP per FADH

Maximum yield per glucose = 38 ATP

• Only achieved by aerobic respiration of mitochondria in eukaryote cells.

• Aerobic respiration by bacteria is less efficient (< 24 ATP).

• Anaerobic respiration is even less efficient.

• Fermentation least efficient (2 ATP)

Hydrolysis of Major Biomolecules

Enyzymes of Hydrolysis: • Proteins by proteases.

• Polysaccharide and other carbohydrates by glycosidase.

•Nucleic acids (DNA or RNA) by nucleases.

• Lipids by lipases.

Amphibolic Nature of Metabolism

Most catabolic pathways have anabolic counterparts, so not all compounds are used to generate ATP, but rather shunted to make new cell biomass.

Energy Source Overview:

• In addition to organisms feeding on organic carbon for energy (chemoorganotrophs).

• There are chemolithotrophs, which gain energy from reduced inorganic compounds (litho = rock).

• There are phototrophs that yield energy from sunlight and do not depend on any chemical energy sources.

• Also note how the terminal (final) electron acceptor determines which respiration type or fermentation.