CEE 370 Lecture #13 10/16/2019 Print version Updated: 16 October 2019 CEE 370 Environmental Engineering Principles Lecture #13 Environmental Biology II Metabolism Reading: Mihelcic & Zimmerman, Chapter 5 Davis & Masten, Chapter 3 David Reckhow CEE 370 L#13 1 Environmental Microbiology Types of Microorganisms Bacteria Viruses Protozoa Rotifers Fungi Metabolism Microbial Disease Microbial Growth 2 CEE 370 L#13 David Reckhow Lecture #13 Dave Reckhow 1
CEE 370 Lecture #13 10/16/2019 Metabolism Energy Production Biosynthesis (Catabolism) (Anabolism) Metabolites (wastes) 3 CEE 370 L#13 David Reckhow An overview of metabolism From: Sawyer, McCarty & Parkin, 1994; also: Sawyer & McCarty, 1978 4 CEE 370 L#13 David Reckhow Lecture #13 Dave Reckhow 2
CEE 370 Lecture #13 10/16/2019 Energy Source Light Chemicals (e.g., glucose) Storage ATP NAD+ Advantages of oxygen as a terminal electron acceptor aerobic anaerobic facultative 5 CEE 370 L#13 David Reckhow ATP 7.3 kcal per mole 6 CEE 370 L#13 David Reckhow Lecture #13 Dave Reckhow 3
CEE 370 Lecture #13 10/16/2019 Nicotinamide Adenine Dinucleotide 7 CEE 370 L#13 David Reckhow Embden-Meyerhof Pathway Glucose 2 ATP Investment 2 ADP Fructose-1,6-diphosphate 2 NAD+ 4 ADP Pay-back 2 NADH 4 ATP 2 Pyruvate Net result: 2 ATPs or 14.6 kcal/mole 8 CEE 370 L#13 David Reckhow Lecture #13 Dave Reckhow 4
CEE 370 Lecture #13 10/16/2019 Advantages of Aerobic Systems If we have aerobic metabolism, rather than fermentation, energy from NADH may be harvested. NADH + H + 3 PO + 3ADP + + 3- 12 O NAD+ + 3ATP + H O 4 2 2 This gives us 6 more ATPs. Then the pyruvate may be further oxidized to carbon dioxide and water, producing 30 more ATPs. The final tally is 38 ATPs or 277 kcal/mole of glucose. 9 CEE 370 L#13 David Reckhow Pathways Generalized view of both aerobic and fermentative pathways Also showing energy transfer ATP NAD From: Sawyer, McCarty & Parkin, 1994; also: Sawyer & McCarty, 1978 10 CEE 370 L#13 David Reckhow Lecture #13 Dave Reckhow 5
CEE 370 Lecture #13 10/16/2019 Metabolic Classification Carbon Source Heterotrophic: other organic matter Autotrophic: inorganic carbon (CO 2 ) Energy Source (electron donor) Chemosynthetic: chemical oxidation Photosynthetic: light energy Terminal Electron Acceptor Aerobic: oxygen Anaerobic: nitrate, sulfate Fermentative: organic compounds 11 CEE 370 L#13 David Reckhow Energy Flow Storage of Energy Photosynthesis Release of Energy Respiration Energy transfers by organisms are inherently inefficient 5-50% capture 12 CEE 370 L#13 David Reckhow Lecture #13 Dave Reckhow 6
CEE 370 Lecture #13 10/16/2019 Aerobic Respiration A Redox reaction Oxidation of Carbon C ( H O ) H O CO 4 H 4 e 2 2 2 Reduction of oxygen or some other terminal electron acceptor O 4 H 4 e 2 H O 2 2 13 CEE 370 L#13 David Reckhow Other TEA: Anaerobic Respiration Nitrate C ( H 2 ) O NO N CO HCO H O 3 2 2 3 2 Manganese 4 2 C ( H 2 ) O Mn Mn CO H O 2 2 Iron 3 2 C ( H 2 ) O Fe Fe CO H O 2 2 Sulfate 2 C ( H 2 ) O SO H S CO H O 4 2 2 2 Fermentation Ecological C ( H 2 ) O CH CO Redox 4 2 methanogenesis Sequence 14 CEE 370 L#13 David Reckhow Lecture #13 Dave Reckhow 7
CEE 370 Lecture #13 10/16/2019 Terminal Electron Acceptors Contribution to Iron the oxidation of Reduction Methano- 1% organic matter genesis Sulfate 23% Reduction Bottom waters of 27% Onondaga Lake, NY (Effler, 1997) Nitrate Reduction 10% Aerobic 39% 15 CEE 370 L#13 David Reckhow Energetics Principles of Gibbs Free Energy and Energy Balance can be applied to microbial growth From: Sawyer, McCarty & Parkin, 1994; also: Sawyer & McCarty, 1978 16 CEE 370 L#13 David Reckhow Lecture #13 Dave Reckhow 8
CEE 370 Lecture #13 10/16/2019 Energetics Cont. Energy Balance Cell synthesis (R c ) Energy (R a ) Electron acceptor (R d ) R f R f R R s c e a d From: Sawyer, McCarty & Parkin, 1994; also: Sawyer & McCarty, 1978 17 CEE 370 L#13 David Reckhow f-values and Yield Portions of electron donor used for: Synthesis (f s ) Energy (f e ) Values are for rapidly growing cells From: Sawyer, McCarty & Parkin, 1994; also: Sawyer & McCarty, 1978 18 CEE 370 L#13 David Reckhow Lecture #13 Dave Reckhow 9
CEE 370 Lecture #13 10/16/2019 Novel Biotransformations Oxidation Toluene dioxygenase (TDO) From: Sawyer, McCarty & Parkin, 1994; also: Sawyer & McCarty, 1978 19 CEE 370 L#13 David Reckhow Overall Types Photoheterotrophs Purple and green non- sulfur bacteria Carbon Source (rare) Heterotrophic Photoautotrophs Cyanobacteria, algae Autotrophic & Plants (primary producers) Energy Source Chemoheterotrophs Most bacteria, fungi, Chemosynthetic (organotrophs) protozoa & animals Chemoautotrophs Photosynthetic Nitrifying, hydrogen , (lithotrophs) iron and sulfur bacteria 20 CEE 370 L#13 David Reckhow Lecture #13 Dave Reckhow 10
CEE 370 Lecture #13 10/16/2019 Enzyme Chemistry Highly dependent on: Temperature pH From: Sawyer, McCarty & Parkin, 1994; also: Sawyer & McCarty, 1978 21 CEE 370 L#13 David Reckhow Enzymatic Reactions Many ways of illustrating the steps Substrate(s) bond to active site Product(s) form via transition state Product(s) are released 22 CEE 370 L#13 David Reckhow Lecture #13 Dave Reckhow 11
CEE 370 Lecture #13 10/16/2019 Note that some Basic Enzyme Kinetics references use k 2 for k -1 , and k 3 for k 2 k 1 Irreversible k 2 → E + S ES E + P ← k- 1 Single intermediate The overall rate is determined by the RLS, k 2 d [ S ] d [ P ] r k 2 ES [ ] dt dt But we don’t know [ES], so we can get it by the SS mass balance d [ ES ] 0 k [ E ][ S ] k [ ES ] k [ ES ] 1 1 2 dt Again, we only know [E o ] or [E tot ], not free [E], so: 0 k [ E ] [ ES ] [ S ] k [ ES ] k [ ES ] 1 o 1 2 23 CEE 370 L#13 David Reckhow Reactants, products and Intermediates Simple Progression of components for simple single intermediate enzyme reaction Shaded block shows steady state intermediates Assumes [S]>>[E] t From Segel, 1975; Enzyme Kinetics 24 CEE 370 L#13 David Reckhow Lecture #13 Dave Reckhow 12
CEE 370 Lecture #13 10/16/2019 Basic Enzyme Kinetics II And solving for [ES], k [ ES ][ S ] k [ ES ] k [ ES ] k [ E ][ S ] 1 1 2 1 o k [ E ][ S ] [ ES ] 1 o k [ S ] k k 1 1 2 [ E ][ S ] [ ES ] o k k [ S ] 1 2 k 1 25 CEE 370 L#13 David Reckhow Michaelis-Menten Irreversible k 1 k 2 → E + S ES E + P ← k- 1 Single intermediate d [ P ] r k 2 ES [ ] dt [ E ][ S ] o [ ES ] k k [ S ] 1 2 k 1 d [ P ] k [ E ][ S ] r [ S ] r 2 o max k k dt K [ S ] [ S ] 1 2 s k 1 26 CEE 370 L#13 David Reckhow Lecture #13 Dave Reckhow 13
CEE 370 Lecture #13 10/16/2019 Michaelis Menten Kinetics Classical substrate plot rmax 100 80 Reaction Rate 60 0.5r max 40 d [ P ] r [ S ] K s max r s dt K [ S ] 20 0 0 20 40 60 80 100 120 27 CEE 370 L#13 David Reckhow Substrate Concentration Maud Menten 28 CEE 370 L#17 David Reckhow Lecture #13 Dave Reckhow 14
CEE 370 Lecture #13 10/16/2019 Substrate and growth d [ P ] d [ S ] 1 dX r If we consider Y dt dt Y dt We can define a microorganism-specific substrate utilization rate, U dX r dt U YX X Y And the maximum rates are then U k max max Y 1 d [ S ] k [ S ] 1 d [ X ] [ S ] max U and s s X dt K [ S ] X dt K [ S ] 29 CEE 370 L#13 David Reckhow Linearizations Lineweaver-Burke Double reciprocal plot Wikipedia version Voet & Voet version CEE 370 L#13 30 David Reckhow Lecture #13 Dave Reckhow 15
CEE 370 Lecture #13 10/16/2019 To next lecture 31 CEE 370 L#13 David Reckhow Lecture #13 Dave Reckhow 16
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