Chapter 3 + some chapter 14 - Microbial Metabolism & Metabolic Diversity of Microbes

Catabolism

• Reactants → products

• Creates ATP from ADP (Energy can be used for anabolism or for movement, transport of nutrients)

• Exergonic reactions: reactions that enable ATP-synthesis (energy is released)

• A reaction that gives off energy has a negative delta G

Anabolism

• Precursors → cellular materials

• Uses ATP

Catabolism

• Reactants → products

• Creates ATP from ADP (Energy can be used for anabolism or for movement, transport of nutrients)

• Exergonic reactions: reactions that enable ATP-synthesis (energy is released)

• A reaction that gives off energy has a negative delta G

Anabolism

• Precursors → cellular materials

• Uses ATP

Exergonic reactions

• Reactions with a negative delta G (spontaneous)

• These enable ATP-synthesis

• ATP is subsequently used for synthesis of cell components

Reducing power

• Is essential for:

  • Energy generation (catabolism)

  • Biosynthesis of cell components (anabolism)

Energy sources for microbes

Metabolic classes of microbes

Reduction potential

• Tendency to donate electrons (so potential to reduce others because is oxidized itself)

• Reduced substance of a redox couple with a more negative E₀ donates electrons to the oxidized substance of a redox couple with a more positive E

What happens when the gap becomes larger in the redox tower?

• More negative which also releases more energy

Redox tower

• Represents the range of possible reduction potentials

• Substances toward the top (so all the negative values) prefer to donate electrons

• Substances toward the bottom (oxidized, so all the positive values) prefer to accept electrons

• The farther the electrons “drop”, the greater the amount of energy released

Reduction potential

• Potential of reducing others (being oxidized itself)

• The tendency to donate electrons (V)

• More negative is better electron donors

• More positive is better electron acceptors

• In redox tower, the electron acceptors are at bottom and donors at the top.

• Molecules that are more neutral (e.g. -0.320) usually bridge the gap between donating and accepting. A good example is NAD/NADH

Electron carriers

• Electrons are never free in the cell, but always bound to a carrier

• Electrons from energy source → electron carrier → terminal electron acceptor (e.g. O₂)

• Cells contain only a limited amount of electron carriers that means that the reduced form must be continuously oxidized (NADH → NAD+), therefore relatively low levels of NAD are needed

• Most common NAD⁺ → NADH

What does the value of G tell us?

• If the reaction is exergonic or endergonic

• Exergonic: Can be a potential energy source for the cell

• Endergonic: Requires an energy input to proceed

• The larger the gap between the half reactions in the redox tower, the more energy and the larger G.

The process of recycling NAD as electron carrier

Method to recycle NAD+/NADH

1. Enzyme I accepts NAD+ and substrate (e- donor)

2. This releases NADH and the rest of the substrate without the electron

3. Now the NADH binds to enzyme II with the substrate (e- acceptor)

4. Afterwards the product has the e- and NADH is turned back into NAD+. The cycle begins again.

What molecules can store this high amount of energy coming from electrons in the short-run

• ATP (most important, for short term processes)

• Pyruvate

• Acetyl-CoA

• Acetyl phosphate

• Glucose 6-phosphate

Lots of these contain phosphate → this is because the carbon phosphate group contains a lot of energy

Long term energy storage

• Involves biosynthesis of insoluble polymers that can be oxidized to generate ATP

• Examples in prokaryotes:

  • Glycogen

  • Poly-B-hydroxybutyrate

  • Polyphosphate

  • Elemental sulfur (S)

Essences of catabolism

1. Substrate-level phosphorylation → ATP synthesis DIRECTLY coupled to an energy generating (exergonic) reaction (Basically ADP → ATP)

2. Electron transport phosphorylation (=oxidative phosphorylation) → ATP synthesis coupled to electron transport, via a respiratory chain. Redox reactions enable ATP synthesis INDIRECTLY

3. Photophosphorylation> light energy used for making ATP

Fermentation

Anaerobic catabolism in which organic compounds donate and accept electrons

• no electron acceptor is available - therefore oxidized intermediates used as electron acceptor

• Only substrate level phosphorylation

• 2 ATP produced → rest of the energy is still present in the products (lactate, ethanol)

Respiration

A metabolic process in which an electron donor is oxidized, transferring electrons through an electron transport chain to a terminal electron acceptor, which may be oxygen (aerobic respiration) or another compound (anaerobic respiration), resulting in ATP production.

Steps of fermentation

1. An organic compound (e.g. glucose) is taken up by the cell

2. The cell oxidizes this compound and forms and energy-rich compound. (To do this NAD+ becomes NADH)

3. This energy-rich compound is used for substrate level phosphorylation where ADP is turned into ATP and that forms an oxidized compound

4. The cell then reduces the compound again and the fermentation product is excreted (also NADH → NAD+)

Glycolysis

1. Investment stage: Start with glucose where two ATP are invested

   • Glucose is separated into two (C3) molecules which each have two phosphate groups (= lots of energy!!)

2. Pay-off stage:

   • NAD⁺ is turned into NADH

   • The C3 molecules are dephosphorylated to form ATP

   • Pyruvate is formed

   • For fermentation:

     • Since we now have NADH which cannot accept any electrons, the pyruvate will receive the electrons

     • The pyruvate can turn into lactate (muscles or certain bacteria) or 2 ethanol and 2 CO₂ products (yeasts)

In total two ATP are invested and four ATP are received back.

What do yeast produce in glycolysis?

• 2 ethanol

• 2 CO₂

What do lactic acid bacteria or your muscles produce?

2 Lactate

Energetic and redox considerations for fermentative microbes

1. Dependent on substrate level phosphorylation → little energy

2. Shortage on external electron acceptors; difficult to reach redox balance

   • This holds especially for strict fermentative microbes.

Besides reduction of intermediates for redox balance, how else can redox be balanced?

By production of H₂

Lactic acid bacteria

• Gram +

• Not sporulating

• Oxygen tolerant (indifferent)

• Lactic acid as main end product

What are the two types of lactic acid bacteria?

• homofermentative: make only lactic acid

• Heterofermentative: Make lactic acid but also ethanol and CO2

  • Heterofermentative will do other pathways besides glycolysis

How is ATP generated in respiration?

• By oxidative phosphorylation

• Drive by the “proton motive force”, which is build up over the cytoplasmic membrane of prokaryotes.

Citric acid cycle

• pathway through which pyruvate is completely oxidized

• the electrons that are released are transferred to NAD+ and FAD

• three intermediates are also precursors for anabolic reactions

1. Start with C6 molecule

2. Then down to C5, to C4

3. C4 combines with C2 to make C6, process starts again.

The electron transport chain

• Complex I - NADH turns into NAD+ and electrons enter the cycle. Protons are pumped across the membrane

• Complex II - FADH2 turns into FAD

• Complex III - Quinones (made in I & II) go into complex III, flow through the cytochromes and eventually through complex IV to the oxygen. Also pumps protons across the membrane

• Complex IV - the electrons from complex III that have gone through the cytochrome go to the oxygen and the complex pumps out two protons.

pH in electron transport chain

• Protons are pumped out and therefore the outside becomes electrically positive and acidic

• The inside becomes electrically negative and alkaline

What is the proton motive force used for?

• ATP production by ATP synthase

• Active transport

• Movement

Electron transport chain in anaerobic respiration

Is possible, instead nitrate is reduced but this will give far less ATP

Order of type of respiration

1. If oxygen is available → it will respire with oxygen

2. If oxygen runs out, it will try to switch to nitrate respiration

3. If nitrate runs out, it will do fermentation because there is no longer an electron acceptor available

Respiration vs. Fermentation

	Respiration	Fermentation
Products	6H2O and 6CO2	2CO2 and 2 Ethanol
ATP	38 ATP	2 ATP
Gibbs free energy	-2844 kJ	-226 kJ

Applications of anabolism

• Making RNA/DNA

• Making fatty acids

How much ATP does glycolysis and CAC produce? (aerobic respiration)

38 ATP

How much ATP does fermentation produce from glucose?

2 ATP per glucose

Flexibility of respiration in bacteria

• Bacteria can often do both aerobic and anaerobic respiration

• E.g. E. coli grows by aerobic respiration (so oxygen is electron acceptor), fermentation (no electron acceptor), or a simple form of anaerobic respiration (with nitrate)

What is the Embden-Meyerhof-Parnas pathway?

Another word for glycolysis

Which intermediate compound(s) in the citric acid cycle is/are often used for biosynthetic pathways as well as carbon catabolism?

Enzymes: α-ketoglutarate, oxaloacetate, and succinyl-CoA

In what form is nitrogen commonly found?

In inorganic forms

Three elements that all microbes need

• Phosphorus

• Selenium

• Sulfur

What class of macromolecules in microbes contributes the most to biomass?

Proteins