Chapter 6 Microbial Metabolism

1. Chapter 6 Microbial Metabolism • Energy Metabolism • Special Metabolism in Microbes • The Relationship between Catabolism and Anabolism • Regulation of Metabolism and Ferment Industry

2. catabolism metabolism anabolism An Overview metabolism metabolism： the sum total of all chemical reactions occurring in the cell catabolism [H] ATP Complex molecules Simple molecules anabolism

3. Section 1 Energy Metabolism in Microbes summarize Chemoheterotroph Organic Compounds ATP Photoheterotroph Primary Energy Sunlight ATP Photoautotroph Inorganic Compounds in Reduced State Chemoautotroph

4. Chemoheterotrophbiological oxidation and energy Release Process——Dehydrogenation, Giving Hydrogen and Accepting Hydrogen (Electron) Function——Releasing Energy (ATP), Engendering Reducing Power [H] and Producing Intermediate Metabolites The breakdown of glucose to pyruvate fermentation respiration（aerobic or anaerobic respiration） biological oxidation

6. 1 The breakdown of glucose to pyruvate Glocose——representative substrate of biological oxidation Embden-Meyerhof-Parnas Pathway (Glycolysis) Hexose Monophosphate Pathway Entner-Doudoroff Pathway (KDPG Pathway) PK (phosphoketolase) pathway

7. Ten steps 1）Embden-Meyerhof-Parnas Pathway(EMP) (Glycolysis, Hexose Diphosphate Pathway) glucosepyruvate with O2：connecte EMP pathway with TCA pathway； without O2：reduce some metabolismproduct, only energy-yielding process. generates ATP by substrate-level phosphorylation： （1） glyceraldehyde 1,3-phosphate 3-phoshoglyceric acid+ ATP； （2） PEP  pyruvate + ATP C6H12O6＋2NAD+＋2ADP＋2Pi→ 2CH3COCOOH＋2NADH＋2H+＋2ATP＋2H2O

8. 2)Hexose Monophosphate Pathway (HMP) (Pentose Phosphate Pathway, Phosphogluconate Pathway, Warburg-Dickens Pathway) • Uses pentoses and NADPH • Operates with glycolysis

9. fructose 6-phosphates： be converted toglucose 6-phosphates ，be returned toPentose Phosphate Pathway glyceraldehyde 3-phosphate： a. through EMP pthway， be converted to pyruvate，intoTCA pthway b. converted toHexose Phosphate, be returned to Pentose Phosphate Pathway The overall reaction： 6 glucose 6-phosphates +12NADP++3H2O → 5 glucose 6-phosphates + 6CO2+12NADPH+12H++Pi

10. 3) Entner-Doudoroff Pathway (KDPG Pathway) 1952,Entner-Doudoroff ：Pseudomonas saccharophila process： （4 septs） 1 Glucoseglucose 6-phosphates 6-phosphogluconate 6-phosphogluconate dehydratase KDPG 2-oxo-3-deoxy-6-phosphogluconate aldolase glyceraldehyde 3-phosphate+ pyruvate Produces NADPH and 1 ATP Does not involve glycolysis Pseudomonas, Rhizobium, Agrobacterium

11. 4) phosphoketolase pathway (PK) a.Pentose phosphoketolas Pathway G xylulose 5-phosphate phosphoketolase acetyl phosphate + glyceraldehyde 3-phosphate pyruvate ethanol Lactic acid 1 G  Lactic acid + ethanol + 1 ATP+ NADPH + H+

12. b.Hexose phosphoketolas Pathway (HK) （Bifidobacterium bifidum） G fructose 6-phosphates phosphoketolase Erythrose-4-P + acetyl-phosphates fructose 6-phosphates acetokinase xylulose -5-P ribulose-5-P Acetic acid phosphoketolase glyceraldehyde 3-phosphate+ acetyl-phosphates Lactic acid Acetic acid 1 G  Lactic acid + 1.5 Acetic acid + 2.5 ATP

13. 2 fermentantion 1. definitions broader ：use microorganism to produce useful metabolic product narrower：under anaerobic conditions,be defined as an energy-yielding process using itself metabolic intermediates as the final hydrogen (electron) accepter organic molecules serve as both electron donors and acceptors. trait： 1） generates ATP by substrate-level phosphorylation； 2）the glucose is partially oxidized，mostly energy in fermention products； 3）lower energy； 4） generates many kinds of fermention products

14. 2. fermentationsorts 1） alcohol fermentation a. yeasts 1 G 2 pyruvate  2 aldehyde + CO2  2 ethanol +2 ATP condiction：pH 3.5~4.5 , without O2 Strain：Saccharomyces cerevisiae, few bacteria(Erwinia amylovora, Sarcine vintriculi) I. Add NaHSO4 NaHSO4 + aldehyde sulfonic hydroxy aldehyde ii.weak basic（pH  7.5） 2 aldehyde  1 acetate + 1 ethanol glycerolfermentation : dihydroxyacetone as hydrogen accepter, hydrolyzed to glycerol （EMP）

15. ethanol+1ATP b. bacteria(Zymomonasmobili, Pseudomonas saccharophila ) homoalcohol fermentation 1 G 2 pyruvate （ED） heteroalcohol fermentation(Thermoanaerobacter ethanolicu) 1 G 2 pyruvate （ Pyruvate formate lyase） formic acids + acetyl-CoA Without Pyruvatedecarboxylate With aldehyde dehydrogenase aldehyde  ethanol

16. 2）Lactic acid fermentation Homolactatefermentation For example, Lactobacillus delbruckii, Streptococcus faecalis —— EMP pathway（pyruvate lactate） Heterolactatefermentation（PK pathway） Leuconostoc mesenteroides（PK） generates energy：1ATP Bifidobacterium bifidum（PK、HK） generates energy： 2G  5 ATP, 1G  2.5ATP

17. Pyruvate formate lyase acetyl-CoA+formyl Methylmalonyl CoA carboxyltransferase oxaloacetate Propionic acid 3) mixed acid, butanediol fermentation a.mixed acid fermentation： ——E.coli, Salmonella , Shiella CO2 + H2 lactate dehydrogenase lactate phosphotransacetylase aldehyde dehydrogenase 1 G pyruvate acetokinase Alcohol dehydrogenase acetate ethanol

18. b.butanediol fermentation ——Enterobacter, Serratia (acetolactate dehydrogenase) pyruvate acetolactate 3-hydroxy butanone （OH-、O2） V.P. test: Red substance diacetyl butanediol neutral Two important reaction: 1. V. P. test 2. methylene red (M.R) test Enterobacter aerogenes: methylene red （-） E.coli: V.P. （-）, methylene red （+）

19. 4) acetone-butanol fermentation （ mixed, acetone:butanol : ethanol = 3：6：1） ——Clostridium acetobutyricum 2 pyruvate 2 acetyl-CoA acetyl-CoA acetyltransferase acetoacetyl-CoA （acetate CoA transferase） （acetoacetate decarboxylase） butanol acetone+CO2

20. 5）stickland reaction Main point：amino acid oxidation couples with other amino acids reduction, generates 1ATP hydrogendonors(oxidation ) amino acid: Ala, Leu, Ile, Val, His, Ser, Phe, Tyr, Try hydrogenacceptors (reduction) amino acid: Gly, Pro, Arg, Met, Leo, and so on.

21. The Stickland reaction is used to oxidize several amino acids: alanine, leucine, isoleucine, valine, phenylalanine, tryptophan, and histidine. Bacteria also ferment amino acids (e.g., alanine, glycine, glutamate, threonine, and arginine) by other mechanisms.

22. NAD+ NAD+ NADH NADH alanine acetate acetate+ ATP -NH3 Oxidized glycine pyruvate acetyl-CoA Reduced alanine -NH3 one amino acid is oxidized and a second amino acid acts as the electron acceptor.

23. 3 Respiration aerobic respiration anaerobic respiration 1. Aerobic Respiration: The final electron acceptor in the electron transport chain is molecular oxygen (O2). Respiration

24. （1）Tricarboxylic Acid Cycle (Krebs Cycle) Give the substrate and products of the tricarboxylic acid cycle. To provide carbon skeletons for use in biosynthesis What chemical intermediate links glycolysis to the TCA cycle? The complete cycle appears to be functional in many aerobic bacteria, free-living protozoa, and most algae and fungi.

25. （2）The Electron Transport Chain Some important electron transport chain carriers of the respiration chain in microbes Nicotianamide adenine dinucleotide (NAD) and nicotianamide adenine dinucleotide phosophate (NADP) Flavin adenine dinucleotide (FAD) and flavin mononucleotide (FMN) Iron-sulphur protein Ubiquinone (Coenzyme Q) Cytochrome system

26. The Mitochondrial Electron Transport Chain. Many of the more important carriers are arranged at approximately the correct reduction potential and sequence. In the eucaryotic mitochondrial are organized into four complexes that are linked by coenzyme Q (I and cytochrome c (Cyt c). Electrons flow from NADH and succinl down the reduction potential gradient to oxygen.

27. The electron transport chain in Prokaryote Main outline: electron accepter multiplicity：O2, NO3-, NO2-, NO-, SO42-, S2-, CO32- et al； Electron donors：H2, S, Fe2+, NH4+, NO2-, G, other orgnisim et al； various cytochrome： a, a1, a2, a4, b, b1, c, c1, c4, c5, d, o； Terminal oxidase： cyt a1, a2, a3, d, o，catalase, peroxid enzyme； Respiration Chain component variable, being branchedrespiration chain： Bacterial chains also may be shorter and have lower P／O ratios than mitochondrial transport chains， from the several position of electron transport chain into and off by Terminal oxidase in several position. E.coli （absent O2） CoQ  cyt.b556  cyt.o cyt.b558  cyt.d The cytochromes 0 branch has moderately high affinity for oxygen，is a proton pump，and operates at higher oxygen concentrations;The cytochromes d branch has very high affinity for oxygen and functions at low oxygen levels。 electron transport multiplicity.

28. （3）Oxidative phosphorylqtion Chemiosmotic hypothesis Chemiosmotic hypothesis-first formulated in 1961 by the British biochemist Peter．Mitchell. According to the Chemiosmotic hypothesis,the electron transport chain is organized so that protons move outward from the mitochondrial matrix and electrons are transported inward. Protons movement may result either from carrier loops,as shown in figure 9.14,or from the action of special proton pumps that derive their energy from electron transport. The result is proton motive force(PMF), composed of a gradient of protons and a membrane porential due to the unequal distribution of charges.When protons return to the mitochondrial matrix driven by the proton motive force,ATP is synthesized in a reversal of the ATP hydrolysis reaction.

29. 2. 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 operations under anaerobic conditions. Nitrate Respiration（ Denitrification） Nitrate reduction outline： a. have completely Respiration system； b. in the absence of O2 only， nitrate reductase A and nitrous acid reductase needing for Denitrification are induced. c. facultativeanaerobic bacteria: Bacillus licheniformis, Paracoccus denitrificans, Pseudomonas aeruginosa and so on.

30. Nitrate Respiration Homotype： NO3- NH3 - N  R - NH2 Hetertype： in the absence of O2 ，use NO3-as the final hydrongen accepter NO3-  NO2  NO   N2O  N2 denitrification： 1) make N（NO3-） in soil reduced into N2，and disappear, reduce edaphic fertility； 2) Denitrification has the importance action in nitrogen cycle. Facultative anaerobe (viz denitrify bacteriua)     NaR NiR NOR N2OR

31. Sulfate Respiration(sulfate reduction） ——in the absent of O2,SO42-, SO32-, S2O32- as the finalelectron accepter outline: a. obligate anaerobic bacteria； b. mostly ancient bacteria c. mostil obligate chemoheterotroph，few mixed； d. end product: H2S； SO42- SO32-  SO2  S  H2S e. use organic nutrients（organic acid, fatty acid, 醇类）as hydrogen donors or electron donors； f. environment： contain SO42- ,anaerobic environment（soil, seawater） For example, Desulfovibrio desulfuricans, D．Gigas, Desulfotomaculum nigrificans and so on.

32. Sulfur Respiration（ Sulfur reduction） ——use element Sulfuras the final electron accepteronly. electron donors：aceticm acid, small peptide, glucose, carbohydrate polymers; For example, Desulfuromonas acetoxidans Carbonate Respiration（ Carbonate reduction ） ——use CO2、HCO3- as the final electron accepter methane-producing bacteria: ——use H2 as electron donors（energy resources), CO2 as accepter，produce CH4; Producing acetic acid bacteria — H2 / CO2 carry out Anaerobic Respiration，product acetic acid

33. other anaerobic respiration ——use Fe3+,Mn2+ , many organic oxide as the final electron accepter fumaric acid succinate + 1 ATP For example, Escherichia, Proteus, Salmonella, Klebsiella in some facultative anaerobic bacteria, Bacteroides, Propionibacterium, Vibrio succinogenes in some anaerobic bacteria. Use Desulfotomaculum auripigmentum reduces AsO43- into As2S3

34. Section 2 special metabolism in microbes 1 Bacterial Photosynthesis 1. Cyclic photophosphorylation 2. Noncyclic photophosphorylation 3. Photosynthesis of purple membrane in halophilic bacteria

35. green sulfur bacteria: Chlorobium green nonsulfur bacteria : Chloroflexus purple sulfur bacteria: Chromatium purple nonsulfur bacteria: Rhodospirillum, Rhodopseudomonas

36. Purple Nonsulfur Bacterial Photosynthesis. Cyclic photophosphorylation The photosynthetic electron transport system in the purple nonsulfur bacterium, Rhodobacter sphaeroides. This scheme is incomplete and tentative. Ubiquinone (Q) is very similar to coenzyme Q. BPh stands bacteriopheophytin. NAD+ and the electron source succinate are in color.

37. Green SulfurBacterial Photosynthesis. Light energy is used to make ATP by cyclic photophosphorylation move electrons from sulfur donors (green and blue) to NAD+ (red). The electron transport chain has a quinone called menaquinone (MK). The photosynthetic electron transport system in the green sulfur bacterium, Chlorobium limicola.

38. 2）noncyelic photophosphorylation Electrons also can travel in a noncyclic pathway involving both photosystems. P700 is excited and donates electrons to ferredoxin as before. In the noncyclic route, however, reduced ferredoxin reduces NADP+ to NADPH . Because the electrons contributed to NADP+ cannot be used to reduce oxidized P700, photosystem II participation is required. It donates electrons to oxidized P700 and generates ATP in the process. The photosystem II antenna absorbs light energy and excites P680, which then reduces pheophytin a. Pheophytin a is chlorophyll a in which two hydrogen atoms have replaced the central magnesium. Electrons subsequently travel to Q (probably a plastoquinone) and down the electron transport chain to P700. Oxidized P680 then obtains an electron from the oxidation of water to O2.

39. noncyelic photophosphorylation

40. Outline : • electrons flow from water all the way to NADP with the aid of energy from two photosystems, • ATP is synthesized by noncyelic photophosphorylation. • one ATP and one NADPH are formed when two electrons travel through noncyclic pathway.

42. 3) Photosynthesis of purple membrane in halophilic bacteria Halobacterium uses bacteriorhodopsin, not chlorophyll, to generate electrons for a chemiosmotic proton pump.

44. 2 Chemolithotroph Biological Oxidation, Energy Release and CO2 Fixation in Chemoautotroph aerobic Oxidation of inorganic electron donors generates ATP by Oxidative Phosphorylation electron donors: H, reducing nitride, reducing sulphide and Fe2＋. Use CO2 Fixation of Calvin cycle as carbon resources CO2 reduced to [CH2O]——consume much energy and reducing power

45. 1. Energy metabolism of nitribacteria Oxidation Nitrobacter: NH3 NO2- Oxidation Nitrosomonas:NO2- NO3- When two genera such Nitrobacter and Nitrosomonas together in a niche, ammonia is grow converted to nitrate, a process called nitrification

46. 2. Hydrogen oxidizing bacteria Main strains: Alcaligenes, Flavobacterium Aquaspirillum Mycobacterium Nocardia and so on. generates energy： 2H2 + O2 — 2 H2O synthesize reaction：2H2 + CO2 — [ CH2O ] + H2O。

48. 3.Sulfur Bacteria Energy Metabolism of Sulfur Bacteria Thiobacillus Energy source: Thiosulfate freely soluble in water and in the neutral condition. The respiratory chain of Thiobacilli: NADH2 dehydrogenase, Fumaric reductase, flacoprotein（FP）, ubliquinone（CoQ）, cyt b, Cytochrome oxidase aa3

49. energy-yielding process : First sept: H2S, S0, S2O32－, oxidatived to SO32－ Second sept: SO32－ oxidatived to SO42－and generates energy

50. 4 iron bacteria and bacterial leaching (1) iron bacteria (iron oxidizing bacteria) oxidative Fe2＋into Fe3 ＋and generates energy. For example, Ferrobacillus, Gallionella,Leptothrix,Crenothrix and Sphaerotilus; Thiobacillus frrooxidans:oxidative, S0 and reducing sulphide, and Fe2＋ oxidatived to Fe3＋，so both Sulfur Bacteria and iron bacteria. mostly obligate Chemoautotrophic, few facultative Chemoautotrophicbacteria, acidophilic bacteria.