1. 3
Microbial Metabolism

2. II. Energetics, Enzymes and Redox
3.3 Energy Classes of Microorganisms
3.4 Bioenergetics
3.6 Electron Donors and Electron Acceptors
3.7 Energy-Rich Compounds

3. 3.3 Energy Classes of Microorganisms
Metabolism
The sum total of all of the chemical reactions that occur in a cell
Catabolic reactions (catabolism)
Energy-releasing metabolic reactions
Anabolic reactions (anabolism)
Biosynthetic metabolic reactions

4. 3.3 Energy Classes of Microorganisms
Microorganisms have a variety of ways to conduct their metabolism
Microorganisms grouped into energy classes
Chemorganotrophs
Chemolithotrophs
Phototrophs

5. Figure 3.5 Metabolic options for conserving energy.

6. 3.3 Energy Classes of Microorganisms
Autotrophs fix (reduce) their own carbon
Heterotrophs rely on carbon fixed (reduced) by another organism

7. 3.4 Bioenergetics
In any chemical reaction, some energy is lost as heat but some energy (Free energy = G): energy released that is available to do work
The change in free energy during a reaction at standard conditions is referred to as ΔG0′
ΔG: free energy that occurs under actual conditions

8. 3.6 Electron Donors and Electron Acceptors
Energy from oxidation–reduction (redox) reactions is used in synthesis of energy-rich compounds (e.g., ATP)
Redox reactions occur in pairs (two half reactions; Figure 3.8)
Electron donor: the substance oxidized in a redox reaction
Electron acceptor: the substance reduced in a redox reaction

9. 3.6 Electron Donors and Electron Acceptors
Half reaction donating e–
Electron
donor
Electron
acceptor
Formation
of water
Net reaction
Half reaction accepting e–
Figure 3.8 Example of an oxidation–reduction reaction.
Energy from oxidation–reduction (redox) reactions is used in synthesis of energy-rich compounds (e.g., ATP)
Figure 3.8

10. 3.6 Electron Donors and Electron Acceptors
Reduction potential (E0′): tendency to donate electrons
Expressed as volts (V)
Substances can be either electron donors or electron acceptors under different circumstances (redox couple)
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 E0′

11. 3.6 Electron Donors and Electron Acceptors
The redox tower represents the range of possible reduction potentials (Figure 3.9)
The reduced substance at the top of the tower donates electrons
The oxidized substance at the bottom of the tower accepts electrons
The farther the electrons "drop," the greater the amount of energy released

12. Figure 3.9 The redox tower.
Figure 3.9

13. 3.6 Electron Donors and Electron Acceptors
Redox reactions usually involve reactions between intermediates (carriers)
Electron carriers are divided into two classes
Prosthetic groups (attached to enzymes)
Example: heme
Coenzymes (diffusible)
Examples: NAD+, NADP

14. 3.7 Energy-Rich Compounds
Chemical energy released in redox reactions is primarily stored in certain phosphorylated compounds (Figure 3.12)
ATP; the prime energy currency
Phosphoenolpyruvate
Glucose 6-phosphate
Chemical energy also stored in coenzyme A, a high energy sulfur compound

15. Anhydride bonds
Ester bond
Ester bond
Anhydride bond
Phosphoenolpyruvate
Adenosine triphosphate (ATP)
Glucose 6-phosphate
Compound
G0′kJ/mol
Thioester
bond
Anhydride bond
ΔG0′< 30kJ
Phosphoenolpyruvate
–51.6
1,3-Bisphosphoglycerate
–52.0
Acetyl phosphate
–44.8
Figure Phosphate bonds in compounds that conserve energy in bacterial metabolism.
ATP
–31.8
ADP
–31.8
Acetyl
Coenzyme A
Acetyl phosphate
Acetyl-CoA
–35.7
Acetyl-CoA
ΔG0′< 30kJ
AMP
–14.2
Glucose 6-phosphate
–13.8
Figure 3.12

16. 3.7 Energy-Rich Compounds
Long-term energy storage involves insoluble polymers that can be oxidized to generate ATP
Examples in prokaryotes
Glycogen
Poly-β-hydroxybutyrate and other polyhydroxyalkanoates
Elemental sulfur
Examples in eukaryotes
Starch
Lipids (simple fats)

17. III. Fermentation and Respiration Overview
3.8 Glycolysis
3.9 Fermentative Diversity and the Respiratory Option
3.10 Respiration: Electron Carriers
3.11 Respiration: The Proton Motive Force
3.12 Respiration: Citric Acid and Glyoxylate Cycle
3.13 Catabolic Diversity

18. III. Fermentation and Respiration Overview
Two key metabolic pathways
Complementary
Overlapping
Definition depends on context-industrial, medical, biochemical

19. Fermentation
In food science fermentation can refer to the production of foods such as yogurt
In chemical engineering it can refer to the production of ethanol as an additive for gasoline
In microbiology it refers to the breakdown of carbon compounds (eg glucose) to smaller compounds with a limited harvest of energy through substrate level phosphorylation and no oxygen used

20. Respiration
In medicine or exercise science respiration refers to breathing
In microbiology respiration refers to the removal of electrons from a substance and their transfer to a terminal acceptor with a significant harvest of energy through oxidative phosphorylation (redox reactions). Oxygen may be used as the terminal acceptor (aerobic respiration) or not (anaerobic respiration).

21. ~ ~ ATP ATP i i Intermediates Energy-rich intermediates P ADP A B B P
Substrate-level phosphorylation
Energized
membrane
ADP + P i
Dissipation of proton
motive force coupled
to ATP synthesis
ATP
Figure 3.13 Energy conservation in fermentation and respiration.
Less energized
membrane
Electron transfer used to generate proton motive force
(b) Oxidative phosphorylation

22. Fermentation: substrate-level phosphorylation; ATP is directly synthesized from an energy-rich intermediate
Respiration: oxidative phosphorylation; ATP is produced from proton motive force formed by transport of electrons

23. Fermentation A basic and important process for microorganisms
A sugar is the starting material and the end product depends on the species
Three Stages (I) Preparation, (II) Energy Harvesting, (III) Reboot

24. Figure 3.14 Fermentation with lactic acid produced
Figure 3.14 Embden–Meyerhof–Parnas pathway (glycolysis).
Figure 3.14

25. 3.8 Glycolysis
Stage I and Stage II (Figure 3.14) called Glycolysis (Embden–Meyerhof pathway): a common pathway for catabolism of glucose
End product of glycolysis is pyruvate (pyruvic acid)
In fermentation pyruvate is processed through Stage III
It accepts electrons so is reduced

26. 3.9 Fermentative Diversity and the Respiratory Option
Fermentations may be classified by products formed
Ethanol
Lactic acid (homolactic vs heterolactic)
Propionic acid
Mixed acids
Butyric acid (extra ATP generated)
Butanol

27. 3.9 Fermentative Diversity and the Respiratory Option
Fermentations may be classified by substrate fermented
Usually NOT glucose
Amino acids
Purines and pyrimidines
Aromatic compounds

28. Disambiguation
Glycolysis aka Emden-Myerhof Pathway or Emden-Myerhof-Parnas Pathway
Entner-Doudoroff Pathway-an alternative way to make pyruvate
Phosphoketolase Pathway-still another way to make pyruvate
NOT pentose phosphate shunt aka phosphogluconate pathway-this one is anabolic

29. Insert pathway name here!

30. 3.10 Respiration: Electron Carriers
In respiration
Electrons are removed from a substance and transferred to one lower on the redox tower.
Compounds called electron carriers move the electrons.
Electrons leave the chain of carriers when they are passed to a terminal electron acceptor
This end product usually exits from the entire system or cell
Carriers stay behind and get reused

31. 3.10 Respiration: Electron Carriers
Important electron carriers include
NADH dehydrogenases
Flavoproteins
Cytochromes (heme)
Iron-sulfur proteins
Quinones

32. 3.10 Respiration: Electron Carriers
Electron are moved by electron transport systems
Membrane-associated
Organize the transfer of electrons
Conserve some of the energy released during transfer as a proton gradient
Use some of the conserved energy to synthesize ATP

33. 3.10 Respiration: Electron Carriers
NADH dehydrogenases: proteins bound to inside surface of cytoplasmic membrane; active site binds NADH and accepts 2 electrons and 2 protons that are passed to flavoproteins
Flavoproteins: contains flavin prosthetic group (e.g., FMN, FAD) that accepts 2 electrons and 2 protons but donates the electrons only to the next protein in the chain (Figure 3.16)

35. E0′ of FMN/FMNH2 (or FAD/FADH2) = –0.22 V
Isoalloxazine ring
Ribitol
Oxidized (FMN)
Figure 3.16 Flavin mononucleotide (FMN), a hydrogen atom carrier.
Reduced (FMNH2)
E0′ of FMN/FMNH2 (or FAD/FADH2) = –0.22 V
Figure 3.16

36. 3.10 Respiration: Electron Carriers
Cytochromes
Proteins that contain heme prosthetic groups (Figure 3.17)
Accept and donate a single electron via the iron atom in heme

37. Figure 3.17 Redox site Porphyrin (Fe2+ Fe3+) ring Heme Cytochrome
Figure 3.17 Cytochrome and its structure.
Cytochrome
Figure 3.17

38. 3.10 Respiration: Electron Carriers
Iron–sulfur proteins
Contain clusters of iron and sulfur (Figure 3.18)
Example: ferredoxin
Reduction potentials vary depending on number and position of Fe and S atoms
Carry electrons

39. Figure 3.18 Arrangement of the iron–sulfur centers of nonheme iron–sulfur proteins.

40. 3.10 Respiration: Electron Carriers
Quinones
Hydrophobic non-protein-containing molecules that participate in electron transport (Figure 3.19)
Accept electrons and protons but pass along electrons only

41. Figure 3.19 Structure of oxidized and reduced forms of coenzyme Q, a quinone.

42. 3.10 Respiration: Electron Carriers
Aerobic respiration
Oxidation using O2 as the terminal electron acceptor
Oxygen is reduced to water-a waste product of respiration
Electron transport generates proton motive force
Proton motive force used to make ATP
Higher ATP yield than fermentations

43. 3.11 Respiration: The Proton Motive Force
Electron transport system oriented in cytoplasmic membrane so that electrons are separated from protons (Figure 3.20)
Electron carriers arranged in membrane in order of their reduction potential
The final carrier in the chain donates the electrons and protons to the terminal electron acceptor

44. Figure 3.20 Generation of the proton motive force during aerobic respiration.

45. 3.11 Respiration: The Proton Motive Force
During electron transfer, several protons are released on outside of the membrane
Protons originate from reduced cofactors (eg NADH) and the spontaneous dissociation of water
Results in generation of pH gradient and an electrochemical potential across the membrane (the proton motive force)
The inside becomes electrically negative and alkaline
The outside becomes electrically positive and acidic

46. 3.11 Respiration: The Proton Motive Force
Complex I (NADH:quinone oxidoreductase)
NADH donates e– to FAD
FADH donates e– to quinone
Complex II (succinate dehydrogenase complex)
Bypasses Complex I
Feeds e– and H+ from FADH directly to quinone pool

47. 3.11 Respiration: The Proton Motive Force
Complex III (cytochrome bc1 complex)
Transfers e– from quinones to cytochrome c
Cytochrome c shuttles e– to cytochromes a and a3
Complex IV (cytochromes a and a3 )
Terminal oxidase; reduces O2 to H2O

48. 3.11 Respiration: The Proton Motive Force
ATP synthase (ATPase): complex that harnesses proton motive force to make ATP; two components (Figure 3.21)
F1: multiprotein extramembrane complex; faces cytoplasm
Fo: proton-conducting intramembrane channel
Reversible; dissipates proton motive force

49. Figure 3.21 c a δ α δ β α ADP + Pi α α β β F1 F1 ATP In In b2 b2 γ γ ε
Figure 3.21 Structure and function of the reversible ATP synthase (ATPase) in Escherichia coli. F0
Membrane F0
c12
Out
Out
Figure 3.21

50. 3.12 Respiration: Citric Acid and Glyoxylate Cycle
Citric acid cycle (CAC): pathway through which pyruvate is completely oxidized to CO2 (Figure 3.22a) (aka Krebs or TCA cycle)
Initial steps (glucose to pyruvate) same as glycolysis
Per glucose molecule, 6 CO2 molecules released and NADH and FADH generated
Plays a key role in catabolism AND biosynthesis
Energetics advantage to aerobic respiration (Figure 3.22b)

51. Figure 3.22a The citric acid cycle.

52. Figure 3.22b The citric acid cycle.

53. 3.12 Respiration: Citric Acid and Glyoxylate Cycle
The citric acid cycle generates many compounds available for biosynthetic purposes
- α-Ketoglutarate and oxaloacetate (OAA): precursors of several amino acids; OAA also converted to phosphoenolpyruvate, a precursor of glucose
Succinyl-CoA: required for synthesis of cytochromes, chlorophyll, and other tetrapyrrole compounds
Acetyl-CoA: necessary for fatty acid biosynthesis

54. 3.12 Respiration: Citric Acid and Glyoxylate Cycle
Organic acids can be metabolized as electron donors and carbon sources by many microbes
C4-C6 citric acid cycle intermediates (e.g., citrate, malate, fumarate, and succinate) are common natural plant and fermentation products and can be readily catabolized through the citric acid cycle alone

55. 3.12 Respiration: Citric Acid and Glyoxylate Cycle
Catabolism of C2-C3 organic acids typically involves production of oxaloacetate through the glyoxylate cycle (Figure 3.23)
A variation of the citric acid cycle
Glyoxylate is a key intermediate

56. Figure 3.23 The glyoxylate cycle.

57. 3.13 Catabolic Diversity
Microorganisms demonstrate a wide range of mechanisms for generating energy (Figure 3.24)
Fermentation
Aerobic respiration
Anaerobic respiration
Chemolithotrophy
Phototrophy

58. 3.13 Catabolic Diversity Anaerobic respiration
The use of electron acceptors other than oxygen
Examples include nitrate (NO3–), ferric iron (Fe3+), sulfate (SO42–), carbonate (CO32–), certain organic compounds
Less energy released compared to aerobic respiration
Dependent on electron transport, generation of a proton motive force, and ATPase activity

59. 3.13 Catabolic Diversity Chemolithotrophy
Uses inorganic chemicals as electron donors
Examples include hydrogen sulfide (H2S), hydrogen gas (H2), ferrous iron (Fe2+), ammonia (NH3)
Typically aerobic
Begins with oxidation of inorganic electron donor
Uses electron transport chain and proton motive force
Autotrophic; uses CO2 as carbon source

60. Figure 3.24 Figure 3.24 Catabolic diversity. Electron donor
(organic compound)
Fermentation
Electron transport/
generation of pmf
Aerobic
respiration
Electron
acceptors
Organic e–
acceptors
Anaerobic respiration
Chemoorganotrophy
Chemotrophs
Photoheterotrophy
Photoautotrophy
Light
Organic
compound
Phototrophs
Electrons from
H2O (oxygenic)
H2S (anoxygenic)
Electron transport/
generation of pmf
Electron
transport
Electron
acceptors
Figure 3.24 Catabolic diversity.
Aerobic respiration
Generation of pmf
Cell material
Cell material
Anaerobic respiration
Chemolithotrophy
Phototrophy
Figure 3.24

61. 3.13 Catabolic Diversity Phototrophy: uses light as energy source
Photophosphorylation: light-mediated ATP synthesis
Photoautotrophs: use ATP for assimilation of CO2 for biosynthesis
Photoheterotrophs: use ATP for assimilation of organic carbon for biosynthesis

62. 3.17 Nitrogen Fixation
Living systems require nitrogen in the form of NH3 or R-NH2
“Fixed” or “reduced” nitrogen, not N2
Only some prokaryotes can fix atmospheric nitrogen

63. 3.17 Nitrogen Fixation
Some nitrogen fixers are free-living, and others are symbiotic
Cyanobacteria are free-living
nitrogen fixers
Soybean root nodules contain
endosymbiotic
Bradyrhizobium japonicum

64. 3.17 Nitrogen Fixation Energetically expensive (8 ATP per N atom)
Requires electron donor, often pyruvate
Reaction is catalyzed by nitrogenase
Sensitive to the presence of oxygen
Fe plus various metal cofactors
Can catalyze a variety of reactions

65. Writing answers-exam questions
Three parts
One: the answer itself (yes, no, etc.)
Two: the reason why this is your answer (“because”)
Three: the examples to back up your reason (Example One shows this, Example Two shows that…)