Micrococcus luteus on blood agar

A microbiologists view of the periodic table Group 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 Period 1 Key: Essential for all microorganisms Essential cations and anions for most microorganisms Trace metals, some essential for some microororganisms 2 Used for special functions Unessential, but metabolized 3 Unessential, not metabolized 4 5 6

Colony morphology , A form of multicellularity?

Streaking for singles. Looking for single colony forming units Isolated colonies at end of streak Confluent growth at beginning of streak

Enzymes lower activation energy no enzyme Substrates (A  B) Activation energy with enzyme Free energy ∆G0 = Gf0(C  D) Gf0(A  B) Products (C  D) Progress of the reaction

Enzymes are recycled Substrate Glyceraldehyde-3-P Dihydroxyacetone-P Fructose 1,6-bisphosphate Products Active site Enzyme–substrate complex Free aldolase Free aldolase

Enzymes are specific for their substrates 3 dimensional structure determined by folding is dependent on side chain interactions determined by charge and hydrophobicity.

ReDox - gaining electrons = reduction losing electrons = oxidation Leo the lion goes Gerrrrrr Electron donor Electron acceptor Electron-donating half reaction Electron-accepting half reaction Formation of water Net reaction

Some ReDox potentials of ETC Redox couple E0 (V) -0.60 -0.50 -0.40 -0.30 (1) -0.20 -0.10 0.0 +0.10 (2) +0.20 +0.30 +0.40 +0.50 +0.60 +0.70 (3) +0.80 +0.90 (1) H2  fumarate2 succinate2 ∆G0  = –86 kJ (2) H2  NO3 NO2 + H2O ∆G0  = –163 kJ (3) H2  O2 H2O ∆G0  = –237 kJ

Fig. 5-10-1 0.0 Redox couple ∆G0  = –86 kJ E0 (V) -0.60 -0.50 -0.40 -0.30 (1) -0.20 -0.10 0.0 +0.10 (1) H2  fumarate2 succinate2 ∆G0  = –86 kJ

Fig. 5-11 NADH  H Oxidized Reduced NAD Nicotinamide Ribose Ribose Adenine Phosphate added in NADP

Enzyme I reacts with electron donor and oxidized form of Fig. 5-12 Reaction 1. Enzyme I reacts with electron donor and oxidized form of coenzyme, NAD+. Reaction 2. Enzyme II reacts with electron acceptor and reduced form of coenzyme, NADH. NAD+ binding site Active site NADH binding site Active site Enzyme I Enzyme II NAD+ Electron donor NADH Electron acceptor Enzyme substrate complex NADH Electron donor oxidized NAD+ Electron acceptor reduced

Bond energies of some important compounds Anhydride bonds Ester bond Ester bond Anhydride bond Phosphoenolpyruvate Adenosine triphosphate (ATP) Glucose 6-phosphate Anhydride bond Thioester bond Acetyl Coenzyme A Acetyl-CoA Acetyl phosphate

Adenosine triphosphate (ATP) Fig. 5-13-1 Anhydride bonds Ester bond Anhydride bond Phosphoenolpyruvate Adenosine triphosphate (ATP) Anhydride bond Thioester bond Acetyl Coenzyme A Acetyl-CoA Acetyl phosphate

Using SLP to drive thermodynamically unfavorable reactions Intermediates in the biochemical pathway Energy-rich intermediates Substrate-level phosphorylation Energized membrane Less energized membrane Oxidative phosphorylation

You must use energy to free energy STAGE I: PREPARATORY REACTIONS Isomerase Glucose Hexokinase Phosphofructokinase Glucose-6- Fructose-6- Fructose-1,6- Aldolase STAGE II: MAKING ATP AND PYRUVATE 2 Glyceraldehyde-3- Glyceraldehyde-3-P dehydrogenase 2 Electrons 2 NAD+ 2 1,3-Bisphosphoglycerate 2 NADH To Stage III Phosphoglycerokinase 2 3-Phosphoglycerate 2 2-Phosphoglycerate Enolase 2 Phosphoenolpyruvate STAGE III: MAKING FERMENTATION PRODUCTS Pyruvate kinase 2 Pyruvate NADH Lactate dehydrogenase Pyruvate decarboxylase Pyruvate:Formate lyase To Stage II NAD+ Acetate formate Formate hydrogenlyase Lactate Acetaldehyde CO2 H2  CO2 NADH Alcohol dehydrogenase NAD+ To Stage II Ethanol

Fig. 5-15-1 Glucose STAGE I: PREPARATORY REACTIONS Glucose-6- Hexokinase Isomerase Phosphofructokinase Glucose-6- Fructose-6- Fructose-1,6-

Investment and return on investment Aldolase STAGE II: MAKING ATP AND PYRUVATE 2 Glyceraldehyde-3- Glyceraldehyde-3-P dehydrogenase 2 Electrons 2 NAD+ 2 1,3-Bisphosphoglycerate 2 NADH To Stage III Phosphoglycerokinase 2 3-Phosphoglycerate 2 2-Phosphoglycerate Enolase 2 Phosphoenolpyruvate

Fig. 5-15-3 Pyruvate 2 STAGE III: MAKING FERMENTATION PRODUCTS NADH Pyruvate kinase 2 Pyruvate NADH Lactate dehydrogenase Pyruvate decarboxylase Pyruvate:Formate lyase To Stage II NAD+ Acetate formate Formate hydrogenlyase Lactate Acetaldehyde CO2 H2  CO2 NADH Alcohol dehydrogenase NAD+ To Stage II Ethanol

Iron-sulfur clusters : a motif for electron transfer R-Cysteine Cysteine-R R-Cysteine Cysteine-R R Cysteine R Cysteine R Cysteine Cysteine R

Fig. 5-20 E0 (V) E0 (V) Complex I –0.22 0.0 Complex II Fumarate Succinate CYTOPLASM 0.1 Complex III Complex IV 0.36 0.39 ENVIRONMENT E0 (V)

F1/Fo ATP synthase and the proton gradient chemiosmosis F1 In b2 Membrane Fo C12 Out

The Balance sheet: The bottom line Pyruvate (three carbons) Key Energetics Balance Sheet for Aerobic Respiration C2 Acetyl-CoA (1) Glycolysis: Glucose  2NAD  2 ATP 2 Pyruvate  4 ATP  2 NADH C4  4 ADP C5 to CAC to Complex I C6 (a) Substrate-level phosphorylation 2 ADP  Pi 2 ATP 8 ATP Oxalacetate2 Citrate3 (b) Oxidative phosphorylation 2 NADH 6 ATP Aconitate3 Malate2 (2) CAC: Pyruvate  4 NAD  GDP  FAD 3 CO2  4 NADH  FADH GTP Isocitrate3 to Complex I to Complex II (a) Substrate-level phosphorylation Fumarate2 1 GDP  Pi 1 GTP 1 GTP  1 ADP 1 ATP  1 GDP 15 ATP ( 2) (b) Oxidative phosphorylation 4 NADH 1 FADH Succinate2 12 ATP 2 ATP –Ketoglutarate2 Succinyl-CoA (3) Sum: Glycolysis plus CAC 38 ATP per glucose

38 ATP per glucose Energetics Balance Sheet for Aerobic Respiration Fig. 5-22b Energetics Balance Sheet for Aerobic Respiration (1) Glycolysis: Glucose  2NAD  2 ATP 2 Pyruvate  4 ATP 2 NADH Key  4 ADP C2 to CAC to Complex I C4 (a) Substrate-level phosphorylation C5 2 ADP  Pi 2 ATP 8 ATP C6 (b) Oxidative phosphorylation 2 NADH 6 ATP (2) CAC: Pyruvate  4 NAD  GDP  FAD 3 CO2  4 NADH  FADH  GTP to Complex I to Complex II (a) Substrate-level phosphorylation 1 GDP  Pi 1 GTP 1 GTP  1 ADP 1 ATP  1 GDP 15 ATP ( 2) (b) Oxidative phosphorylation 4 NADH 1 FADH 12 ATP 2 ATP 38 ATP per glucose (3) Sum: Glycolysis plus CAC

Fig. 5-23 Chemoorganotrophy Chemolithotrophy Phototrophy Fermentation Organic compound CO2 Carbon flow in respirations Electron transport/ Proton motive force Biosynthesis O2 Electron acceptors S0 NO3– SO42 Organic e– acceptors Aerobic respiration Anaerobic respiration Chemoorganotrophy Inorganic compound CO2 Electron transport/ Proton motive force Carbon flow Electron acceptors S0 O2 NO3– SO42 Biosynthesis Chemolithotrophy Photoheterotrophy Light Photoautotrophy Organic compound Electron transport CO2 Carbon flow Carbon flow Proton motive force Biosynthesis Biosynthesis Phototrophy

Chemoorganotrophy Chemolithotrophy Electron transport/ Fig. 5-23ab Fermentation Organic compound CO2 Carbon flow in respirations Electron transport/ Proton motive force Biosynthesis O2 Electron acceptors Organic e– acceptors Aerobic respiration S0 NO3– SO42 Anaerobic respiration Chemoorganotrophy Inorganic compound CO2 Electron transport/ Proton motive force Carbon flow Electron acceptors S0 O2 NO3– SO42 Biosynthesis Chemolithotrophy

Phototrophy Photoheterotrophy Light Photoautotrophy Electron transport Fig. 5-23c Photoheterotrophy Light Photoautotrophy Electron transport Organic compound CO2 Carbon flow Carbon flow Proton motive force Biosynthesis Biosynthesis Phototrophy

Glutamate family Proline Glutamine Arginine -Ketoglutarate Fig. 5-25 Glutamate family Proline Glutamine Arginine -Ketoglutarate Citric acid cycle Aspartate family Asparagine Lysine Methionine Threonine Isoleuine Oxalacerate Alanine family Valine Leucine Pyruvate Glycolysis Serine family Glycine Cysteine 3-Phosphoglycerate Phospho- enolpyruvate Aromatic family Phenylalanine Tyrosine Tryptophan Chorismate Erythrose-4-P

Glutamate  Oxalacetate Fig. 5-26 -Ketoglutarate  NH3 Glutamate Glutamate dehydrogenase Glutamate  NH3 Glutamine Glutamine synthetase Glutamate  Oxalacetate -Ketoglutarate  Aspartate Transaminase Glutamine  -Ketoglutarate 2 Glutamate Glutamate synthase

CO2 Amino group of aspartate Glycine Formyl group (from folic acid) Fig. 5-27 CO2 Amino group of aspartate Glycine Formyl group (from folic acid) Formyl group (from folic acid) Amide nitrogen of glutamine Ribose-5-P Inosinic acid NH3 Aspartic acid CO2 Orotic acid Uridylate

Palmitate (16 C) 4 C 6 C 14 C 8 C 12 C 10 C Fig. 5-28 Acetoacetyl-CoA Acetyl-ACP Malonyl-ACP Acetoacetyl-CoA Palmitate (16 C) 4 C 6 C 14 C 8 C 12 C 10 C

Control of pathways: feedback inhibition (noncompetitive inhibition) Starting substrate The allosteric enzyme Enzyme A Intermediate I Enzyme B Intermediate II Feedback inhibition Enzyme C Intermediate III Enzyme D End product

(allosteric effector) Substrate Fig. 5-30 Enzyme Active site Allosteric site End product (allosteric effector) Substrate INHIBITION: Substrate cannot bind; enzyme reaction inhibited ACTIVITY: Enzyme reaction proceeds

Phosphoenol pyruvate Erythrose 4-phosphate Initial substrates  1 2 3 Fig. 5-31 Phosphoenol pyruvate Erythrose 4-phosphate Initial substrates  1 2 3 DAHP synthases (isoenzymes 1, 2, 3) DAHP Chorismate Tyrosine Tryptophan Phenylalanine

Glutamine synthetase, a paradigm of allosteric control 100 Enzyme activity Glutamine concentration Glutamine Relative GS activity Glutamine 50 GS GS–AMP6 GS–AMP12 3 6 9 12 AMP groups added AMP

The makings of a microbe

also

Cofactors galore: Take your vitamins!

Tab. 5-4

Thinking thermo!