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Micrococcus luteus on blood agar
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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
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Colony morphology , A form of multicellularity?
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Streaking for singles. Looking for single colony forming units
Isolated colonies at end of streak Confluent growth at beginning of streak
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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
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Enzymes are recycled Substrate Glyceraldehyde-3-P Dihydroxyacetone-P
Fructose 1,6-bisphosphate Products Active site Enzyme–substrate complex Free aldolase Free aldolase
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Enzymes are specific for their substrates
3 dimensional structure determined by folding is dependent on side chain interactions determined by charge and hydrophobicity.
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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
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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 O H2O ∆G0 = –237 kJ
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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
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Fig. 5-11 NADH H Oxidized Reduced NAD Nicotinamide Ribose Ribose
Adenine Phosphate added in NADP
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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
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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
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Adenosine triphosphate (ATP)
Fig Anhydride bonds Ester bond Anhydride bond Phosphoenolpyruvate Adenosine triphosphate (ATP) Anhydride bond Thioester bond Acetyl Coenzyme A Acetyl-CoA Acetyl phosphate
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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
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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
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Fig. 5-15-1 Glucose STAGE I: PREPARATORY REACTIONS Glucose-6-
Hexokinase Isomerase Phosphofructokinase Glucose-6- Fructose-6- Fructose-1,6-
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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
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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
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Iron-sulfur clusters : a motif for electron transfer
R-Cysteine Cysteine-R R-Cysteine Cysteine-R R Cysteine R Cysteine R Cysteine Cysteine R
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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)
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F1/Fo ATP synthase and the proton gradient
chemiosmosis F1 In b2 Membrane Fo C12 Out
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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
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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
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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
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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
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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
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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
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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
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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
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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
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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
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(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
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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
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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
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The makings of a microbe
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also
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Cofactors galore: Take your vitamins!
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Tab. 5-4
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Thinking thermo!
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