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1Metabolism LecturesOutline:! Part I: Fermentations! Part II: Respiration! Part III: Metabolic DiversityLearning objectives are:! Learn about anaerobic respiratory metabolisms.! How can an inorganic compound be use as an energy source.Respiration Review2NADHQ2NAD+2H+2H+H+H+O24H+2 H2OH+H+percytolow Eo’ electron flow hi Eo’ATPADP3H+Electron Tower2Anaerobic Respiration! Anaerobic metabolism is of clinical importance:– Deep tissue infections can lead to abscess formation,foul-smelling pus, and tissue destruction! Uses inorganic and organic molecules other thanoxygen as terminal electron acceptors–! Extensive list of electron acceptors– oxyanions, metals, metal oxides, organic acids, inorganic! Energy and carbon sources are diverse–Metabolic Classification Based onOxygen Concentrations! Points of reference:– Atmospheric oxygen is ~21% (v/v) (or 2.1 x 105 parts permillion)– Low solubility in water: up to 14 parts per million (T and Pdependent)! Remember metabolic classifications:– Strict aerobe (non-fermentative, respires oxygen)– Strict anaerobe (sensitive to oxygen)– Facultative anaerobe: (fermentative and/or respiration)– Microaerophilic (or microaerophile)! 40:1 anaerobes to facultative anaerobes in humanfeces3Diversity of electron acceptors forrespiration! Organic compounds:– Eg. fumarate, dimethylsulfoxide (DMSO), Trimethylamine-N-oxide (TMAO)! Inorganic compounds:– Eg. NO3-, NO2-, SO42-, S0, SeO42-, AsO43-! Metals:– Eg. Fe3+, Mn4+, Cr6+! Minerals/solids:– Eg. Fe(OH)3, MnO2! Gasses:– Eg. NO, N2O, CO2Why is there so muchdiversity?How can prokaryotesaccomplish this?Dehydrogenase:LactateSuccinateFormateNADHGlycerophosphateHydrogenaseMQUQfumarateCyt b, Fe/S, FADFumarate reductaseCyt b, Fe/S, MoDMSODMSO reductaseCyt b, Fe/S, MoTMAOTMAO reductaseCyt b, Fe/S, MoNO3-Nitrate reductaseElectron donor modules Electron acceptor “modules”Answer:4Modularity of electron transport chainswhat do most of these have in common?Example 1.Nitrate reduction5Figure 24.19 Nitrogen cycleFigure 24.19 Nitrogen cycle78% N2Nitrate reducing bacteriaNitrate reducing bacteria! Contribute to denitrification (removal of ?)! Beneficial process for sewage treatment plantsNitrogenous waste good food for algae! Dissimilatory nitrate reduction widespread in microbes– Used for making energy via oxidative phosphorylation! Nitrate is a strong oxidant similar to oxygen! Some microbes can take Nitrate all the way to Nitrogengas:– Pseudomonas stutzeri– E0’ +0.74 V compared to +0.82 for 1/2O2/H2O– How many electrons are used from NO3- to N2?6Dissimulative NO3- reductionDenitrification by Pseudomonas stutzeri7Denitrification by Pseudomonas stutzeri! Four terminal reductases– Nap: Nitrate reductase (Mo-containing enzyme)– Nir: Nitrite reductase– Nor: Nitric oxide reductase– N2or: Nitrous oxide reductase! All can function independently but they operate in unisonDissimilatory nitrate reduction: Biochemistry! Electron donor: lactate, formate, H2, others– Uses special dehydrogenases for these.! Enzymes are membrane-bound! Periplasmic nitrate reductases (NapA) containsa molybdenum cofactor! Coupled to the generation of PMF! ATP synthesized by oxidative phosphorylation8Nitrate vs. oxygen vs. denitrification respirationoxygennitritedenitrificationHow much energyis made byreducing nitrate tonitrite with NADH?Nitrate= N(x) + 3O2- ! x + 3(-2) = -1 ! x=Nitrite= N(x) + 2O2- ! x + 2(-2) = -1 ! x=Determining oxidation state of N and # of electrons:?e-Nitrate(NO3-)Nitrite(NO2-)?NADH?NAD+example?ATPWe only need to oxidize ______ NADH for this:NADH + H+NAD+ + 2H+ + 2e-What’s reduce and oxidized?Find !Eo’ of nitrate/nitrite and NAD+/NADH Use Nernst Eq to find !Go’9Example 2.Arsenate reductionArsenate " arseniteEo’+0.139 Vhttp://phys4.harvard.edu/~wilsonArsenic respiring bacteria and human health! Arsenic is mainly agroundwater pollutant! Affects ~140 million peopleamong ~70 countries! Arsenate (As[V]):– Like phosphate: H2AsO4-– Affects ATP synthesis! Arsenite (As[III]): H3AsO3– More toxic than As(V)– Binds proteins– Causes DNA damage! Microbes respire arsenateand make arsenite! Medical Geology problem10Shewanella sp.strain ANA-32As(V)2As(III)As2S3LactateAcetate+CO2Dividing cellIsolation of strainRespiring O2Respiring AsO43-Example 3. Iron112Fe(III)2Fe(II)NADH2 NAD+c-hem eQH2QCMOMFeIII-oxideIron oxidereducingbacteria! Examples:– Geobacter, Shewanella,Rhodoferrax! How do they do it?Chemolithotrophy and Oxidation ofInorganic MoleculesA. A pathway used by a small number of microorganismscalled chemolithotrophsB. Produces a significant but low yield of ATPC. The electron acceptor is commonly O2, some othersinclude sulfate and nitrateD. The most common electron donors are hydrogen,reduced nitrogen compounds, reduced sulfurcompounds, and ferrous iron (Fe2+)12Geological, biological, and anthropogenic sources of reduced inorganiccompounds supporting chemolithotrophsTypical habitats of chemolithotrohs:Near the interface ofoxic/anoxic conditionsEnergy yields from various inorganicelectron donors:13Oxidation of sulfur-compounds! E.g.: Sulfur oxidizing Thiobacillii– Thiobacillus thiooxidans Thiobacillus ferrooxidans! Produces sulfuric acid (H2SO4)– Acidification of soil– Dissolution of minerals, e.g. CaCO3Reactions !G˚’ kJ/mol H2S + 2 O2 >>> SO42- + 2H+ -798.2 HS- + 1/2O2 + H+ >>> S0 + H2O -209 S0 + H2O + 1.5 O2 >>> SO42- + 2H+ -587 S2O32- + H2O + 2O2 >>> 2 SO42- + 2H+ -818 Lecture Summary! Anaerobic respiration– Alternative terminal electron acceptors are used– ATP generated by oxidative phosphorylation– Often not as energetically favorable as oxygen respiration– Anaerobic electron transport chains are branched– Ecologically and medically significant– In some cases toxic metals are used as electron acceptors! Chemolithotrophy– Energy sources are reduced inorganic compounds– Chemolithotrophs often live near redox gradients where thereis a mixture of reduced and oxidized


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