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Chapter 5 Microbial Metabolism Lectures prepared by Christine L. Case Copyright © 2010 Pearson Education, Inc. Q&A All E. coli look alike through a microscope; so how can E. coli O157 be differentiated? Copyright © 2010 Pearson Education, Inc. Learning Objectives After completing this chapter, you should be able to: Distinguish catabolism from anabolism and how they interact during metabolism Identify the role of ATP Identify the components of an enzyme Describe the mechanism of an enzymatic reaction List factors that influence enzyme activity Distinguish between competitive and noncompetitive inhibition Explain oxidation, reduction, and redox Discuss why enzymatic reactions are important Explain the overall function of metabolic pathways List the steps involved in aerobic carbohydrate catabolism and identify major products of each step List the steps involved in anaerobic respiration and identify major products of each step List some of the major products of fermentation List the steps involved in photosynthesis Compare and contrast the light-dependent reactions versus the light-independent reactions of photosynthesis Categorize the various nutritional patterns among organisms according to carbon source and mechanisms of carbohydrate catabolism and ATP generation Describe the major types of anabolism and their relationship to catabolism Copyright © 2010 Pearson Education, Inc. Catabolic and Anabolic Reactions Metabolism: The sum of the chemical reactions in an organism Copyright © 2010 Pearson Education, Inc. Catabolic and Anabolic Reactions Catabolism: Provides energy and building blocks for anabolism. Anabolism: Uses energy and building blocks to build large molecules Copyright © 2010 Pearson Education, Inc. Role of ATP in Coupling Reactions Figure 5.1 Copyright © 2010 Pearson Education, Inc. Catabolic and Anabolic Reactions A metabolic pathway is a sequence of enzymatically catalyzed chemical reactions in a cell Metabolic pathways are determined by enzymes Enzymes are encoded by genes ANIMATION Metabolism: Overview Copyright © 2010 Pearson Education, Inc. Collision Theory The collision theory states that chemical reactions can occur when atoms, ions, and molecules collide Activation energy is needed to disrupt electronic configurations Reaction rate is the frequency of collisions with enough energy to bring about a reaction. Reaction rate can be increased by enzymes or by increasing temperature or pressure Copyright © 2010 Pearson Education, Inc. Energy Requirements of a Chemical Reaction Copyright © 2010 Pearson Education, Inc. Figure 5.2 Enzyme Components Biological catalysts Specific for a chemical reaction; not used up in that reaction Apoenzyme: Protein Cofactor: Nonprotein component Coenzyme: Organic cofactor Holoenzyme: Apoenzyme plus cofactor Copyright © 2010 Pearson Education, Inc. Components of a Holoenzyme Copyright © 2010 Pearson Education, Inc. Figure 5.3 Important Coenzymes NAD+ NADP+ FAD Coenzyme A Copyright © 2010 Pearson Education, Inc. Enzyme Specificity and Efficiency The turnover number is generally 1 to 10,000 molecules per second. This is the number of substrate molecules that are converted every second. ANIMATION Enzymes: Overview ANIMATION Enzymes: Steps in a Reaction Copyright © 2010 Pearson Education, Inc. The Mechanism of Enzymatic Action Copyright © 2010 Pearson Education, Inc. Figure 5.4a Enzyme Classification Oxidoreductase: Oxidation-reduction reactions Transferase: Transfer functional groups Hydrolase: Hydrolysis Lyase: Removal of atoms without hydrolysis Isomerase: Rearrangement of atoms Ligase: Joining of molecules, uses ATP Copyright © 2010 Pearson Education, Inc. Factors Influencing Enzyme Activity Temperature pH Substrate concentration Inhibitors Copyright © 2010 Pearson Education, Inc. Factors Influencing Enzyme Activity Temperature and pH denature proteins Copyright © 2010 Pearson Education, Inc. Figure 5.6 Effect of Temperature on Enzyme Activity Copyright © 2010 Pearson Education, Inc. Figure 5.5a Effect of pH on Enzyme Activity Copyright © 2010 Pearson Education, Inc. Figure 5.5b Effect of Substrate Concentration on Enzyme Activity Copyright © 2010 Pearson Education, Inc. Figure 5.5c Enzyme Inhibitors: Competitive Inhibition Copyright © 2010 Pearson Education, Inc. Figure 5.7a–b Enzyme Inhibitors: Competitive Inhibition ANIMATION Competitive Inhibition Copyright © 2010 Pearson Education, Inc. Enzyme Inhibitors: Noncompetitive Inhibition ANIMATION Non-competitive Inhibition Copyright © 2010 Pearson Education, Inc. Figure 5.7a, c Enzyme Inhibitors: Feedback Inhibition Copyright © 2010 Pearson Education, Inc. Figure 5.8 Ribozymes RNA that cuts and splices RNA Copyright © 2010 Pearson Education, Inc. Oxidation-Reduction Reactions Oxidation: Removal of electrons Reduction: Gain of electrons Redox reaction: An oxidation reaction paired with a reduction reaction Copyright © 2010 Pearson Education, Inc. Oxidation-Reduction Copyright © 2010 Pearson Education, Inc. Figure 5.9 Oxidation-Reduction Reactions In biological systems, the electrons are often associated with hydrogen atoms. Biological oxidations are often dehydrogenations. ANIMATION Oxidation-Reduction Reactions Copyright © 2010 Pearson Education, Inc. Representative Biological Oxidation Copyright © 2010 Pearson Education, Inc. Figure 5.10 The Generation of ATP ATP is generated by the phosphorylation of ADP Copyright © 2010 Pearson Education, Inc. Substrate-Level Phosphorylation Energy from the transfer of a high-energy PO4– to ADP generates ATP Copyright © 2010 Pearson Education, Inc. Oxidative Phosphorylation Energy released from transfer of electrons (oxidation) of one compound to another (reduction) is used to generate ATP in the electron transport chain Copyright © 2010 Pearson Education, Inc. Photophosphorylation Light causes chlorophyll to give up electrons. Energy released from transfer of electrons (oxidation) of chlorophyll through a system of carrier molecules is used to generate ATP. Copyright © 2010 Pearson Education, Inc. Metabolic Pathways of Energy Production Copyright © 2010 Pearson Education, Inc. Carbohydrate Catabolism The breakdown of carbohydrates to release energy Glycolysis Krebs cycle Electron transport chain Copyright © 2010 Pearson Education, Inc. Glycolysis The oxidation of glucose to pyruvic acid produces ATP and NADH Copyright © 2010 Pearson Education, Inc. Figure 5.11 Preparatory Stage of Glycolysis 2 ATP are used Glucose is split to form 2 glucose-3-phosphate Copyright © 2010 Pearson Education, Inc. Figure 5.12, steps 1–5 Energy-Conserving Stage of Glycolysis 2 glucose-3-phosphate oxidized to 2 pyruvic acid 4 ATP produced 2 NADH produced Copyright © 2010 Pearson Education, Inc. Figure 5.12, steps 6–10 Glycolysis Glucose + 2 ATP + 2 ADP + 2 PO4– + 2 NAD+ 2 pyruvic acid + 4 ATP + 2 NADH + 2H+ ANIMATION Glycolysis: Overview ANIMATION Glycolysis: Steps Copyright © 2010 Pearson Education, Inc. Alternatives to Glycolysis Pentose phosphate pathway Uses pentoses and NADPH Operates with glycolysis Entner-Doudoroff pathway Produces NADPH and ATP Does not involve glycolysis Pseudomonas, Rhizobium, Agrobacterium Copyright © 2010 Pearson Education, Inc. Cellular Respiration Oxidation of molecules liberates electrons for an electron transport chain ATP is generated by oxidative phosphorylation Copyright © 2010 Pearson Education, Inc. Intermediate Step Pyruvic acid (from glycolysis) is oxidized and decarboyxlated Copyright © 2010 Pearson Education, Inc. Figure 5.13 The Krebs Cycle Oxidation of acetyl CoA produces NADH and FADH2 ANIMATION Krebs Cycle: Overview ANIMATION Krebs Cycle: Steps Copyright © 2010 Pearson Education, Inc. The Krebs Cycle Copyright © 2010 Pearson Education, Inc. Figure 5.13 The Electron Transport Chain A series of carrier molecules that are, in turn, oxidized and reduced as electrons are passed down the chain Energy released can be used to produce ATP by chemiosmosis ANIMATION Electron Transport Chain: Overview Copyright © 2010 Pearson Education, Inc. Chemiosmotic Generation of ATP Copyright © 2010 Pearson Education, Inc. Figure 5.16 An Overview of Chemiosmosis ANIMATION Electron Transport Chain: The Process Copyright © 2010 Pearson Education, Inc. Figure 5.15 A Summary of Respiration Aerobic respiration: The final electron acceptor in the electron transport chain is molecular oxygen (O2). Anaerobic respiration: The final electron acceptor in the electron transport chain is not O2. Yields less energy than aerobic respiration because only part of the Krebs cycles operates under anaerobic conditions. Copyright © 2010 Pearson Education, Inc. Respiration ANIMATION Electron Transport Chain: Factors Affecting ATP Yield Copyright © 2010 Pearson Education, Inc. Figure 5.16 Anaerobic Respiration Electron Acceptor Products NO3– NO2–, N2 + H2O SO4– H2S + H2O CO32 – CH4 + H2O Copyright © 2010 Pearson Education, Inc. Carbohydrate Catabolism Pathway Eukaryote Prokaryote Glycolysis Cytoplasm Cytoplasm Intermediate step Cytoplasm Cytoplasm Krebs cycle Mitochondrial matrix Cytoplasm ETC Mitochondrial inner membrane Plasma membrane (cristae) Copyright © 2010 Pearson Education, Inc. Carbohydrate Catabolism Energy produced from complete oxidation of one glucose using aerobic respiration ATP Produced NADH Produced FADH2 Produced Glycolysis 2 2 0 Intermediate step 0 2 Krebs cycle 2 6 2 Total 4 10 2 Pathway Copyright © 2010 Pearson Education, Inc. Carbohydrate Catabolism ATP produced from complete oxidation of one glucose using aerobic respiration Pathway By Substrate-Level Phosphorylation By Oxidative Phosphorylation From NADH From FADH 0 Glycolysis 2 6 Intermediate step 0 6 Krebs cycle 2 18 4 Total 4 30 4 Copyright © 2010 Pearson Education, Inc. Carbohydrate Catabolism 36 ATPs are produced in eukaryotes Really, 38 are produced, but 2 were required for glycolysis, so net gain is 36 ATPs Pathway By Substrate-Level Phosphorylation By Oxidative Phosphorylation From NADH From FADH 0 Glycolysis 2 6 Intermediate step 0 6 Krebs cycle 2 18 4 Total 4 30 4 Copyright © 2010 Pearson Education, Inc. Fermentation Any spoilage of food by microorganisms (general use) Any process that produces alcoholic beverages or acidic dairy products (general use) Any large-scale microbial process occurring with or without air (common definition used in industry) Copyright © 2010 Pearson Education, Inc. Fermentation Scientific definition: Releases energy from oxidation of organic molecules Does not require oxygen Does not use the Krebs cycle or ETC Uses an organic molecule as the final electron acceptor Copyright © 2010 Pearson Education, Inc. An Overview of Fermentation ANIMATION Fermentation Copyright © 2010 Pearson Education, Inc. Figure 5.18a End-Products of Fermentation Copyright © 2010 Pearson Education, Inc. Figure 5.18b Fermentation Alcohol fermentation: Produces ethanol + CO2 Lactic acid fermentation: Produces lactic acid Homolactic fermentation: Produces lactic acid only Heterolactic fermentation: Produces lactic acid and other compounds Copyright © 2010 Pearson Education, Inc. Types of Fermentation Copyright © 2010 Pearson Education, Inc. Figure 5.19 A Fermentation Test Copyright © 2010 Pearson Education, Inc. Figure 5.23 Types of Fermentation Copyright © 2010 Pearson Education, Inc. Table 5.4 Types of Fermentation Copyright © 2010 Pearson Education, Inc. Table 5.4 Lipid Catabolism Copyright © 2010 Pearson Education, Inc. Figure 5.20 Catabolism of Organic Food Molecules Copyright © 2010 Pearson Education, Inc. Figure 5.21 Protein Catabolism Protein Extracellular proteases Deamination, decarboxylation, dehydrogenation, desulfurylation Copyright © 2010 Pearson Education, Inc. Amino acids Organic acid Krebs cycle Protein Catabolism Decarboxylation Copyright © 2010 Pearson Education, Inc. Figure 5.22 Protein Catabolism Desulfurylation Copyright © 2010 Pearson Education, Inc. Figure 5.24 Protein Catabolism Urea Urease Copyright © 2010 Pearson Education, Inc. NH3 + CO2 Clinical Focus Figure B Biochemical Tests Used to identify bacteria. Copyright © 2010 Pearson Education, Inc. Clinical Focus Figure A Photosynthesis Copyright © 2010 Pearson Education, Inc. Figure 4.15 Photosynthesis Photo: Conversion of light energy into chemical energy (ATP) Light-dependent (light) reactions Synthesis: Carbon fixation: Fixing carbon into organic molecules Light-independent (dark) reaction: Calvin-Benson cycle ANIMATION Photosynthesis: Overview Copyright © 2010 Pearson Education, Inc. Photosynthesis Oxygenic: 6 CO2 + 12 H2O + Light energy C6H12O6 + 6 H2O + 6 O2 Anoxygenic: 6 CO2 + 12 H2S + Light energy C6H12O6 + 6 H2O + 12 S ANIMATION: Photosynthesis: Comparing Prokaryotes and Eukaryotes Copyright © 2010 Pearson Education, Inc. Cyclic Photophosphorylation ANIMATION Photosynthesis: Light Reaction: Cyclic Photophosphorylation Copyright © 2010 Pearson Education, Inc. Figure 5.25a Noncyclic Photophosphorylation ANIMATION Photosynthesis: Light Reaction: Noncyclic Photophosphorylation Copyright © 2010 Pearson Education, Inc. Figure 5.25b Calvin-Benson Cycle ANIMATION Photosynthesis: Light Independent Reactions Copyright © 2010 Pearson Education, Inc. Figure 5.26 Photosynthesis Compared Copyright © 2010 Pearson Education, Inc. Table 5.6 Chemotrophs Use energy from chemicals Chemoheterotroph Glucose NAD+ ETC Pyruvic acid NADH Energy is used in anabolism Copyright © 2010 Pearson Education, Inc. ADP + P ATP Chemotrophs Use energy from chemicals Chemoautotroph, Thiobacillus ferrooxidans 2Fe2+ NAD+ ETC 2Fe3+ NADH ADP + P ATP 2 H+ Energy used in the Calvin-Benson cycle to fix CO2 Copyright © 2010 Pearson Education, Inc. Phototrophs Use light energy Chlorophyll ETC Chlorophyll oxidized ADP + P ATP Photoautotrophs use energy in the Calvin-Benson cycle to fix CO2 Photoheterotrophs use energy Copyright © 2010 Pearson Education, Inc. Requirements of ATP Production Copyright © 2010 Pearson Education, Inc. Figure 5.27 A Nutritional Classification of Organisms Copyright © 2010 Pearson Education, Inc. Figure 5.28 A Nutritional Classification of Organisms Copyright © 2010 Pearson Education, Inc. Figure 5.28 A Nutritional Classification of Organisms Copyright © 2010 Pearson Education, Inc. Figure 5.28 Metabolic Diversity among Organisms Nutritional Type Energy Source Carbon Source Example Photoautotroph Light CO2 Oxygenic: Cyanobacteria plants Anoxygenic: Green, purple bacteria Photoheterotroph Light Organic compounds Green, purple nonsulfur bacteria Chemoautotroph Chemical CO2 Iron-oxidizing bacteria Chemoheterotroph Chemical Organic compounds Fermentative bacteria Animals, protozoa, fungi, bacteria. Chemolithotroph Inorganic chemicals CO2 Sulfur oxidizing bacteria, Nitrifying bacteria, Iron bacteria, Hydrogen bacteria Copyright © 2010 Pearson Education, Inc. Check Your Understanding Check Your Understanding Almost all medically important microbes belong to which of the aforementioned groups? 5-23 Copyright © 2010 Pearson Education, Inc. Metabolic Pathways of Energy Use Learning Objective 5-24 Describe the major types of anabolism and their relationship to catabolism. ANIMATION Metabolism: The Big Picture Copyright © 2010 Pearson Education, Inc. Polysaccharide Biosynthesis Copyright © 2010 Pearson Education, Inc. Figure 5.29 Lipid Biosynthesis Copyright © 2010 Pearson Education, Inc. Figure 5.30 Pathways of Amino Acid Biosynthesis Copyright © 2010 Pearson Education, Inc. Figure 5.31a Amino Acid Biosynthesis Copyright © 2010 Pearson Education, Inc. Figure 5.31b Purine and Pyrimidine Biosynthesis Copyright © 2010 Pearson Education, Inc. Figure 5.32 The Integration of Metabolism Amphibolic pathways: Metabolic pathways that have both catabolic and anabolic functions Copyright © 2010 Pearson Education, Inc. Amphibolic Pathways Copyright © 2010 Pearson Education, Inc. Figure 5.33 Amphibolic Pathways Copyright © 2010 Pearson Education, Inc. Figure 5.33