North Carolina State University Chapter 5 Microbial Metabolism Basic Chemical Reactions Underlying Metabolism Metabolism Collection of controlled biochemical reactions that take place within a microbe Ultimate function of metabolism is to reproduce the organism 2012 Pearson Education Inc. Basic Chemical Reactions Underlying Metabolism Metabolic Processes Guided by Eight Elementary Statements Every cell acquires nutrients Metabolism requires energy from light or from catabolism of nutrients Energy is stored in adenosine triphosphate (ATP) Cells catabolize nutrients to form precursor metabolites Precursor metabolites, energy from ATP, and enzymes are used in anabolic reactions Enzymes plus ATP form macromolecules Cells grow by assembling macromolecules Cells reproduce once they have doubled in size 2012 Pearson Education Inc. Basic Chemical Reactions Underlying Metabolism 2012 Pearson Education Inc. ANIMATION Metabolism: Overview Basic Chemical Reactions Underlying Metabolism Catabolism and Anabolism Two major classes of metabolic reactions Catabolic pathways Break larger molecules into smaller products Exergonic Anabolic pathways Synthesize large molecules from the products of catabolism Endergonic 2012 Pearson Education Inc. Figure 5.1 Metabolism Energy lost as heat Nutrients Energy stored Energy used Energy lost as heat Precursor molecules Larger building blocks Energy storage (carbohydrates, lipids, etc.) Macromolecules Cellular processes (cell growth, cell division, etc.) Cellular structures (membranes, ribosomes, etc.) ANABOLISM Basic Chemical Reactions Underlying Metabolism Oxidation and Reduction Reactions Electron transfer from an electron donor to an electron acceptor Reactions always occur simultaneously Cells use electron carriers to carry electrons (often in H atoms) Three important electron carriers Nicotinamide adenine dinucleotide (NAD + ) Nicotinamide adenine dinucleotide phosphate (NADP + ) Flavine adenine dinucleotide (FAD) FADH 2
2012 Pearson Education Inc. Figure 5.2 Oxidation-reduction, or redox, reactions Electron donor Electron acceptor Oxidation Reduction Oxidized donor Reduced acceptor Basic Chemical Reactions Underlying Metabolism 2012 Pearson Education Inc. ANIMATION Oxidation-Reduction Reactions Basic Chemical Reactions Underlying Metabolism ATP Production and Energy Storage Organisms release energy from nutrients Stored in high-energy phosphate bonds (ATP) Phosphorylation organic phosphate is added to substrate Cells phosphorylate ADP to ATP in three ways Substrate-level phosphorylation Oxidative phosphorylation Photophosphorylation Anabolic pathways use some energy by breaking phosphate bonds 2012 Pearson Education Inc. Basic Chemical Reactions Underlying Metabolism The Roles of Enzymes in Metabolism Enzymes are organic catalysts Increase likelihood of a reaction Six categories of enzymes based on mode of action Hydrolases Isomerases Ligases or polymerases Lyases Oxidoreductases Transferases 2012 Pearson Education Inc. Basic Chemical Reactions Underlying Metabolism 2012 Pearson Education Inc. ANIMATION Enzymes: Overview Basic Chemical Reactions Underlying Metabolism The Roles of Enzymes in Metabolism Makeup of enzymes Many protein enzymes are complete in themselves Apoenzymes are inactive if not bound to nonprotein cofactors Binding of apoenzyme and its cofactor(s) yields holoenzyme Some are RNA molecules called ribozymes 2012 Pearson Education Inc. Figure 5.3 Makeup of a protein enzyme Inorganic cofactor Active site Holoenzyme Coenzyme (organic cofactor) Apoenzyme (protein) Basic Chemical Reactions Underlying Metabolism The Roles of Enzymes in Metabolism Enzyme activity Enzymes lower the activation energy Enzyme-substrate specificity Active site complementary to shape of the substrate 2012 Pearson Education Inc. Figure 5.4 Effect of enzymes on chemical reactions Reactants E n e r g y
Progress of reaction Products Activation energy without enzyme Activation energy with enzyme Figure 5.5 Enzymes fitted to substrates-overview Figure 5.6 The process of enzymatic activity (Fructose 1,6-bisphosphate) Substrate Enzyme Enzyme- substrate complex Products (Fructose 1,6- bisphosphate aldolase) Glyceraldehyde-3P Dihydroxyacetone-P Basic Chemical Reactions Underlying Metabolism 2012 Pearson Education Inc. ANIMATION Enzymes: Steps in a Reaction Basic Chemical Reactions Underlying Metabolism The Roles of Enzymes in Metabolism Enzyme activity Many factors influence the rate of enzymatic reactions Temperature pH Enzyme and substrate concentrations Presence of inhibitors Inhibitors Substances that block an enzymes active site Do not denature enzymes Three types 2012 Pearson Education Inc. Figure 5.7 Effects of temperature, pH, and substrate concentration on enzyme activity-overview Figure 5.8 Denaturation of protein enzymes Functional protein Denatured protein Figure 5.9 Competitive inhibition of enzyme activity-overview Basic Chemical Reactions Underlying Metabolism 2012 Pearson Education Inc. ANIMATION Enzymes-Substrate Interaction: Competitive Inhibition Figure 5.10 Allosteric control of enzyme activity-overview Basic Chemical Reactions Underlying Metabolism 2012 Pearson Education Inc. ANIMATION Enzyme-Substrate Interaction: Noncompetitive Inhibition Figure 5.11 Feedback inhibition Feedback inhibition Bound end-product (allosteric inhibitor) Substrate Enzyme 1 Allosteric site Intermediate A Intermediate B Enzyme 2 Enzyme 3 End-product Pathway shuts down Pathway operates Carbohydrate Catabolism Carbohydrate Catabolism Many organisms oxidize carbohydrates as primary energy source for anabolic reactions Glucose most common carbohydrate used Glucose catabolized by two processes: cellular respiration and fermentation 2012 Pearson Education Inc. Figure 5.12 Summary of glucose catabolism Acetyl-CoA Pyruvic acid (or derivative) Formation of fermentation end-products 2 Pyruvic acid Glucose G L Y C O L Y S I S Respiration Fermentation KREBS CYCLE Electrons Carbohydrate Catabolism Glycolysis Occurs in cytoplasm of most cells Involves splitting of a six-carbon glucose into two three-carbon sugar molecules Substrate-level phosphorylation: direct transfer of phosphate between two substrates Net gain of two ATP molecules, two molecules of NADH, and precursor metabolite pyruvic acid 2012 Pearson Education Inc. Carbohydrate Catabolism 2012 Pearson Education Inc. ANIMATION Glycolysis: Overview Carbohydrate Catabolism Glycolysis Divided into three stages involving 10 total steps Energy-investment stage Lysis stage Energy-conserving stage 2012 Pearson Education Inc. Figure 5.13 Glycolysis-overview Carbohydrate Catabolism 2012 Pearson Education Inc. ANIMATION Glycolysis: Steps Figure 5.14 Example of substrate-level phosphorylation Holoenzyme Phosphoenolpyruvate (PEP) Pyruvic acid Phosphorylation Carbohydrate Catabolism Cellular Respiration Resultant pyruvic acid completely oxidized to produce ATP by series of redox reactions Three stages of cellular respiration 1. Synthesis of acetyl-CoA 2. Krebs cycle 3. Final series of redox reactions (electron transport chain) 2012 Pearson Education Inc. Figure 5.15 Formation of acetyl-CoA Pyruvic acid Decarboxylation Acetate Coenzyme A Acetyl-coenzyme A (acetyl-CoA) Respiration Fermentation Carbohydrate Catabolism Cellular Respiration Synthesis of acetyl-CoA Results in Two molecules of acetyl-CoA Two molecules of CO 2
Two molecules of NADH 2012 Pearson Education Inc. Carbohydrate Catabolism Cellular Respiration The Krebs cycle Great amount of energy remains in bonds of acetyl-CoA Transfers much of this energy to coenzymes NAD + and FAD Occurs in cytosol of prokaryotes and in matrix of mitochondria in eukaryotes 2012 Pearson Education Inc. Carbohydrate Catabolism Cellular Respiration The Krebs cycle Six types of reactions in Krebs cycle Anabolism of citric acid Isomerization reactions Hydration reaction Redox reactions Decarboxylations Substrate-level phosphorylation 2012 Pearson Education Inc. Figure 5.16 The Krebs cycle Fermentation Respiration Acetyl-CoA Oxaloacetic acid Malic acid Fumaric acid Succinic acid Succinyl-CoA -Ketoglutaric acid Isocitric acid KREBS CYCLE Citric acid OOH OOH OOH OOH OOH HOO OOH OOH OOH OOH OOH OOH OOH OOH OOH OOH OOH Carbohydrate Catabolism 2012 Pearson Education Inc. ANIMATION Krebs Cycle: Overview Carbohydrate Catabolism 2012 Pearson Education Inc. ANIMATION Krebs Cycle: Steps Carbohydrate Catabolism Cellular Respiration The Krebs cycle Results in Two molecules of ATP Two molecules of FADH 2
Six molecules of NADH Four molecules of CO 2 2012 Pearson Education Inc. Carbohydrate Catabolism Cellular Respiration Electron transport Most significant ATP production occurs from electron transport chain (ETC) Carrier molecules pass electrons from one to another to final electron acceptor Energy from electrons used to pump protons (H + ) across the membrane, establishing a proton gradient Located in cristae of eukaryotes and in cytoplasmic membrane of prokaryotes 2012 Pearson Education Inc. Figure 5.17 An electron transport chain Path of electrons FMN Oxidized Reduced Reduced Oxidized FeS Oxidized Oxidized Reduced Reduced CoQ Cyt Cyt Cyt Reduced Reduced Oxidized Oxidized Final electron acceptor 2 2 2 2 Fermentation Respiration Carbohydrate Catabolism 2012 Pearson Education Inc. ANIMATION Electron Transport Chain: Overview Carbohydrate Catabolism Cellular Respiration Electron transport Four categories of carrier molecules Flavoproteins Ubiquinones Metal-containing proteins Cytochromes Aerobic respiration: oxygen serves as final electron acceptor Anaerobic respiration: molecule other than oxygen serves as final electron acceptor 2012 Pearson Education Inc. Figure 5.18 One possible arrangement of an electron transport chain Bacterium Exterior Cytoplasmic membrane Cytoplasm Intermembrane space Matrix Mitochondrion Exterior of prokaryote or intermembrane space of mitochondrion FMN Phospholipid membrane NADH from glycolysis, Krebs cycle, pentose phosphate pathway, and Entner-Doudoroff pathway FADH 2
from Krebs cycle Ubiquinone Cytoplasm of prokaryote or matrix of mitochondrion Cyt c 2
Cyt b Cyt c Cyt a Cyt a 3 ATP synthase
Carbohydrate Catabolism 2012 Pearson Education Inc. ANIMATION Electron Transport Chain: Process Carbohydrate Catabolism 2012 Pearson Education Inc. ANIMATION Electron Transport Chain: Factors Affecting ATP Yield Carbohydrate Catabolism Cellular Respiration Chemiosmosis Use of electrochemical gradients to generate ATP Create proton gradient from energy released in redox reactions of ETC Protons flow down electrochemical gradient through ATP synthases that phosphorylate ADP to ATP Called oxidative phosphorylation because proton gradient created by oxidation of components of ETC ~34 ATP molecules formed from one molecule of glucose 2012 Pearson Education Inc. Carbohydrate Catabolism Alternatives to Glycolysis Yield fewer molecules of ATP than glycolysis Reduce coenzymes and yield different metabolites needed in anabolic pathways Two pathways Pentose phosphate pathway Entner-Doudoroff pathway 2012 Pearson Education Inc. Figure 5.19 Pentose phosphate pathway Glucose Glucose 6-phosphate Glucose 6-phosphogluconic acid Pentose phosphate sugars Ribulose t-phosphate Xylulose 5-phosphate Ribose 5-phosphate Sedoheptulose 7-phosphate Glyceraldehyde 3-phosphate (G3P) Erythrose 4-phosphate Erythrose 6-phosphate Glucose 6-phosphate Glyceraldehyde 3-phosphate (G3P) To synthesis of amino acids To step 1 of glycolysis or reenter pentose phosphate pathway To step 6 of glycolysis To step 2 of glycolysis To step 6 of glycolysis To synthesis of nucleotides To Calvin-Benson cycle of photosynthesis To anabolic reactions requiring electron donors Figure 5.20 Entner-Douoroff pathway Glucose Glucose 6-phosphate 6-Phosphogluconic acid 2-Keto-3-deoxy- 6-phosphogluconic acid Glyceraldehyde 3-phosphate (G3P) Steps 610 of glycolysis Pyruvic acid Pyruvic acid To Kerb cycle or fermentation Carbohydrate Catabolism Fermentation Sometimes cells cannot completely oxidize glucose by cellular respiration Cells require constant source of NAD +
Cannot be obtained simply using glycolysis and Krebs cycle Fermentation pathways provide cells with source of NAD +
Partial oxidation of sugar or other metabolites to release energy Uses organic molecule within cell as final electron acceptor 2012 Pearson Education Inc. Figure 5.21 Fermentation Respiration Fermentation Pyruvic acid Lactic acid Acetaldehyde Ethanol Figure 5.22 Representative fermentation products and the organisms that produce them Glucose Pyruvic acid Organisms Fermentation Fermentation products CO 2 , propionic acid Lactic acid CO 2 , ethanol Acetone, isopropanol Swiss cheese Cheddar cheese, yogurt, soy sauce Wine, beer Nail polis remover, rubbing alcohol Propionibacterium Aspergillus Lactobacillus Streptococcus Saccharomyces Clostridium Carbohydrate Catabolism 2012 Pearson Education Inc. ANIMATION Fermentation Other Catabolic Pathways Lipid Catabolism Protein Catabolism 2012 Pearson Education Inc. Figure 5.23 Catabolism of a fat molecule-overview Figure 5.24 Protein catabolism Polypeptide Proteases Extracellular fluid Amino acids Cytoplasmic membrane Cytoplasm Deamination To Krebs cycle Photosynthesis Many organisms synthesize organic molecules from inorganic carbon dioxide Capture light energy and use it to synthesize carbohydrates from CO 2 and H 2 O by a process called photosynthesis 2012 Pearson Education Inc. Photosynthesis 2012 Pearson Education Inc. ANIMATION Photosynthesis: Overview Photosynthesis Chemicals and Structures Chlorophylls Important to organisms that capture light energy with pigment molecules Composed of hydrocarbon tail attached to light- absorbing active site centered on magnesium ion Active sites similar to cytochrome molecules in ETC Structural differences cause absorption at different wavelengths 2012 Pearson Education Inc. Photosynthesis Chemicals and Structures Photosystems Arrangement of molecules of chlorophyll and other pigments to form light-harvesting matrices Embedded in cellular membranes called thylakoids In prokaryotes invagination of cytoplasmic membrane In eukaryotes formed from inner membrane of chloroplasts Arranged in stacks called grana Stroma is space between outer membrane of grana and thylakoid membrane 2012 Pearson Education Inc. Figure 5.25 Photosynthetic structures in a prokaryote-overview Photosynthesis Chemicals and Structures Two types of photosystems Photosystem I (PS I) Photosystem II (PS II) Photosystems absorb light energy and use redox reactions to store energy in the form of ATP and NADPH Light-dependent reactions depend on light energy Light-independent reactions synthesize glucose from carbon dioxide and water 2012 Pearson Education Inc. Photosynthesis Light-Dependent Reactions As electrons move down the chain, their energy is used to pump protons across the membrane Photophosphorylation uses proton motive force to generate ATP Photophosphorylation can be cyclic or noncyclic 2012 Pearson Education Inc. Figure 5.26 Reaction center of a photosystem Light Acceptor Reaction center Reaction center chlorophyll Possible path of energy transfer Photosystem I Figure 5.27 Light-dependent reactions of photosynthesis: Cyclic and noncyclic phosphorylation-overview Photosynthesis 2012 Pearson Education Inc. ANIMATION Photosynthesis: Light Reaction: Cyclic Photophosphorylation Photosynthesis 2012 Pearson Education Inc. ANIMATION Photosynthesis: Light Reaction: Noncyclic Photophosphorylation Photosynthesis Light-Independent Reactions Do not require light directly Use ATP and NADPH generated by light- dependent reactions Key reaction is carbon fixation by Calvin-Benson cycle Three steps Fixation of CO 2 Reduction Regeneration of RuBP 2012 Pearson Education Inc. Figure 5.28 Simplified diagram of the Calvin-Benson cycle CALVIN-BENSON CYCLE Ribulose bisphosphate (RuBP) 3-Phosphoglyceric acid G3P 6 3 3 3 O 2
3 5 1 6 6 6 6 6 6 6 1,3-Bisphosphoglyceric acid Glyceraldehyde 3-phosphate (G3P) Glucose 6-phosphate G3P G3P Glucose From the Calvin-Benson cycle or glycolysis From light- dependent reactions of photosynthesis or catabolic pathways Photosynthesis 2012 Pearson Education Inc. ANIMATION Photosynthesis: Light-Independent Reaction Other Anabolic Pathways 2012 Pearson Education Inc. Anabolic reactions are synthesis reactions requiring energy and a source of metabolites Energy derived from ATP from catabolic reactions Many anabolic pathways are the reverse of catabolic pathways Reactions that can proceed in either direction are amphibolic
Figure 5.29 Role of gluconeogenesis in the biosynthesis of complex carbohydrates Glucose Starch, celluose Glycogen Peptidoglycan Glycerol (from fat) Fatty acids (from fat) Amino acids (from protein) CALVIN- BENSON CYCLE Glucose 6-phosphate Fructose 6-phosphate Fructose 1,6-bisphosphate G3P DHAP 2 2 Oxaloacetic acid Pyruvic acid Acetyl-CoA O 2
G L U C O N E O G E N E S I S Figure 5.30 Biosynthesis of fat, a lipid Fats Glycerol Fatty acids DHAP CALVIN- BENSON CYCLE Glyceraldehyde 3-phosphate (G3P) Glycolysis Acetyl-CoA Reverse of beta-oxidation Figure 5.31 Synthesis of amino acids via amination and transamination-overview Figure 5.32 Biosynthesis of nucleotides DNA and RNA Pyrimidine nucleotides Purine nucleotides PABA Folic acid (vitamin in humans) Photosynthesis Aspartic acid (from Krebs cycle) Glutamine (derived from glutamic acid from Krebs cycle) Glucose 6-phosphate PENTOSE PHOSPHATE PATHWAY Ribose 5-phosphate Glycolysis Phosphoglyceric acid Glycine Integration and Regulation of Metabolic Function Cells synthesize or degrade channel and transport proteins Cells often synthesize enzymes needed to catabolize a substrate only when substrate is available If two energy sources are available, cells catabolize the more energy-efficient of the two first Cells synthesize metabolites they need, cease synthesis if metabolite is available 2012 Pearson Education Inc. Integration and Regulation of Metabolic Function Eukaryotic cells isolate enzymes of different metabolic pathways within membrane- bounded organelles Cells use allosteric sites on enzymes to control activity of enzymes Feedback inhibition slows/stops anabolic pathways when product is in abundance Cells regulate amphibolic pathways by requiring different coenzymes for each pathway 2012 Pearson Education Inc. Integration and Regulation of Metabolic Function Two types of regulatory mechanisms Control of gene expression Cells control amount and timing of protein (enzyme) production Control of metabolic expression Cells control activity of proteins (enzymes) once produced 2012 Pearson Education Inc. Figure 5.33 Integration of cellular metabolism (shown in an aerobic organism) Proteins METABOLIC PATHWAYS FOR THE POLYMERIZATION OF MACROMOLECULES Nucleic acids Amino acids Other sugars Nucleotides Lipids Polysaccharides PENTOSE PHOSPHATE PATHWAY ATP AND PRECURSOR METABOLIC PATHWAYS GLYCOLYSIS GLUCONEOGENESIS Fructose 1,6-bisphosphate Glucose Glucose 6-phosphate Glyceraldehyde 3-phosphate (G3P) Glycerol Fatty acids 3-Phosphoglyceric acid Pyruvic acid Acetyl-CoA NH 3
O 2
KREBS CYCLE CALVIN- BENSON CYCLE Photosynthetic organisms INTERMEDIATE METABOLIC PATHWAYS KEY: Catabolic pathway Anabolic pathway Light Integration and Regulation of Metabolic Function 2012 Pearson Education Inc. ANIMATION Metabolism: The Big Picture