349 Metabolism IV: VI. Anaerobic respiration VII. Chemolithotrophy VIII. Anabolism

350 Reoxidation of reduced electron carriers by a process analogous to aerobic respiration, but using a terminal electron acceptor other than O 2. VI. Anaerobic respiration PMF is formed and ATP is synthesized by electron transport phosphorylation. Used by microbes capable of anaerobic respiration when O 2 is not available. TB

351 A. Anaerobic respiration external terminal electron acceptor is not O 2 eg. NO 3 - (nitrate), Fe 3 +, SO 4 -, CO 2, CO 3 2-, fumarate or another organic molecule O2O2

352 Growth substrates Oxidized products Oxidized electron carriers Reduced electron carriers fumarate NO 3 - SO 4 2- CO 2 succinate NO 2 -, N 2 H 2 S CH 4 PMF various electron transport chains

Nitrate reduction NO 3 - NO 2 - a form of anaerobic respiration in which NO 3 - is the terminal electron acceptor nitrate reductase used by Escherichia coli and some other microorganisms when O 2 is absent

354 NO 3 - denitrification 2. Denitrification reduction of nitrate all the way to N 2 through anaerobic respiration Important in agriculture and sewage treatment N2N2 gas

Respiration with sulfur or sulfate SO 4 2- H2SH2S elemental sulfur or SO 4 2- is the terminal electron acceptor S0S0 H2SH2S reduction smelly gases

356 B. Less free energy is released in anaerobic respiration than in aerobic respiration Oxidized form / Reduced form Reduction potential Eo' (Volts) CO 2 / glucose (C 6 H 12 O 2 )(- 0.43) 2 H + / H 2 (- 0.42) NAD+ / NADH(- 0.32) SO 4 2- / H 2 S(- 0.22) pyruvate / lactate(- 0.19) O 2 / H 2 O(+ 0.82) fumarate / succinate(+ 0.03) NO 3 - / NO 2 -(+ 0.42)

357 VII. Chemolithotrophy Use of inorganic compounds as the energy source (primary electron donor) Many chemolithotrophs use O 2 as the terminal electron acceptor H 2 + 1/2 O 2 H 2 O

358 A. Examples of chemolithotrophs H 2 hydrogen-oxidizing bacteria H 2 Ssulfide-oxidizing bacteria Fe 2+ iron-oxidizing bacteria NH 3 ammonia-oxidizing bacteria (NH 3  NO 2 - ) NO 2 - nitrite-oxidizing bacteria (NO 2 -  NO 3 - )

Example of chemolithotrophy: aerobic sulfide (H 2 S) oxidation H 2 S + 2 O 2 inorganic electron donor Boiling sulfur pot, Yellowstone National Park  SO H +

360 Ammonia oxidizer NH 3  NO 2 - Nitrite oxidizer NO 2 -  NO Examples of chemolithotrophy: ammonia oxidation and nitrite oxidation

361 B. Possible metabolic strategies for generating energy on early earth anaerobic chemolithotrophy fermentation anaerobic respiration anoxygenic photosynthesis

362 H2H2 ADP + Pi ATP Cytoplasmic membrane In Out A hypothetical primitive energy- generating system on early earth primitive ATPase primitive hydrogenase Proton motive force (PMF) 2 H + 2 e - inorganic electron acceptor (not O 2 )

363 VIII. Anabolism (Biosynthesis) Nutrients Macromolecules and other cell components Anabolism Energy Energy source (eg. sugar or H 2 ) Waste Catabolism Energy

364 Cells are made of molecules. Nucleic acids Proteins Polysaccharides Lipids small molecules

365 A. Building cell components requires energy (ATP) reductant (NADPH) C H O N P S a source of carbon a source of nitrogen some P and other nutrients

366 carbon source (organic carbon) heterotrophautotroph (CO 2 ) energy source chemoorganotroph (organic chemical) chemolithotroph (inorganic chemical e.g. H 2 S, H 2, NH 3 ) phototroph (light) B. Classification of organisms according to

367 C. Cell carbon sugars acetyl CoA organic acids CO 2 autotrophs NH 3 amino acids protein fatty acids lipid nucleotides nucleic acids P, NH 3 Cell carbon: organic carbon source (e.g. glucose) glycolysis, TCA heterotrophs

368 D. Sugar / polysaccharide metabolism Sugars are needed for polysaccharides (cell wall, glycogen) nucleic acids (DNA, RNA) O O hexosespentoses small molecules (ATP, NAD(P) + cAMP, coenzymes, etc.)

UDP-glucose is a precursor to polysaccharides and peptidoglycan. O  — P-O-  O - O  O= P-O  O - HOCH 2 O CH 2 OH O N O NH O (don't memorize structure) UDP = uridine diphosphate

Gluconeogenesis A pathway for making glucose-6-P from noncarbohydrate sources (e.g. acids from TCA).

Gluconeogenesis is the reversal of glycolysis starting with PEP, but with a few different enzymes. glucose-6-P PEP CO 2 pyruvate TCA OAA succinate gluconeogenesis

Pentose phosphate pathway a. makes pentoses (ribulose-5-P) from the decarboxylation of glucose-6-P b. also makes NADPH for biosynthetic reactions

Deoxyribonucleotides for DNA are made from the reduction of the 2'- hydroxyl of ribonucleotides. OCH 2 HOH O N NH 2 N N N O P P P OCH 2 OH O N NH 2 N N N O P P P NADPH NADP + ATP deoxy- ATP

374 ribose-5-P ribonucleotides  RNA deoxyribo- nucleotides  DNA glucose  glucose-6-P  glucose-1-P  UDP-glucose (uridine diphosphoglucose) ribulose-5-P Sugar summary Gluconeogenesis TCA  PEP  OAA glycolysis UTP polysaccharides peptidoglycan, cell walls pentose phosphate pathway NADPH NADP + pyruvate

375 E. Amino acid biosynthesis 1. Requires an acid (carbon skeleton) and an amino group O C – OH H 2 N – C – H R carboxylic acid amino group

Some carbon skeletons are made in glycolysis and the TCA cycle 5 main amino acid precursors a.  -ketoglutarate (5C) b. oxaloacetate (4C) c. pyruvate (3C) d. phosphoglycerate (3C) e. PEP (3C), (erythrose-4-P)

377 Carbon skeletons for amino acids PEP CO 2 pyruvate TCA OAA (glucose) (acCoA)  -KG phosphoglycerate

378 O C - O - O = C CH 2 COO -  -ketoglutarate O C - O - H 3 N - C - H CH 2 COO - glutamate + NH 3 NADPH NADP + 3. The amino group for glutamate can come directly from ammonia.

379 O C - O - O = C CH 2 COO -  -ketoglutarateglutamate 4. The amino group for most other amino acids comes from glutamate through transamination (amino transfer). oxaloacetate (OAA) aspartate O C - O - H 3 N - C - H CH 2 COO - +

380 F. Purine and pyrimidine biosynthesis is very complex. 1. The carbons and nitrogens come from amino acids, NH 3, CO 2, and formyl (HCOO-) groups. N N N N C C from formyl attached to folic acid * *

Folic acid carries the formyl groups in purine biosynthesis. 3. Sulfanilamide is a "growth factor analog" that inhibits purine biosynthesis by inhibiting the production of folic acid.

382 D. Fatty acids 1. In general, saturated fatty acids are built two carbons at a time from acetyl CoA. CH 3 C~SCoA O (8) ATP, NADPH COO- palmitic acid

Unsaturated fatty acids have 1 or more cis -double bonds increase fluidity of membranes COO-

Acetyl CoA and succinyl CoA and play important roles in anabolism. acetyl CoA  fatty acid biosynthesis succinyl CoA  heme biosynthesis

385 Study objectives 1. Understand anaerobic respiration and the examples presented in class. Define nitrate reduction, denitrification, sulfate reduction. 2. Understand chemolithotrophy and the examples presented in class. 3. Examples of integrative questions: Compare and contrast aerobic respiration, anaerobic respiration, chemolithotrophy, and fermentation. Given the description of a catabolic strategy, be prepared to identify the type of metabolism being used. Contrast sulfate reduction and sulfide oxidation. 4. Be able to classify microorganisms based on energy source and carbon source. 5. Understand the roles of glycolysis and the TCA cycle in the synthesis of cellular macromolecules. 6. What type of polymers are synthesized from UDP-glucose? 7. What are the functions of gluconeogenesis and the pentose phosphate pathway? 8. How are deoxyribonucleotides for DNA made from ribonucleotides?

Know the sources of carbon and nitrogen for amino acid biosynthesis. How are amino groups transferred to acids to make amino acids? 11. Understand the role of folic acid in nucleotide biosynthesis. 12. How does sulfanilamide inhibit the growth of microorganisms? 13. Humans do not make their own folates. Why is the drug sulfanilamide toxic to certain microorganisms but not to humans? 14. Know the anabolic roles of acetyl CoA and succinyl CoA as described in class.