Metabolism Lectures. Outline:! Part I: Fermentations! Part II: Respiration! Part III: Metabolic Diversity

Transcription

1 Outline:! Part I: Fermentations! Part II: Respiration! Part III: Metabolic Diversity Metabolism Lectures Learning objectives are:! Learn about anaerobic respiratory metabolisms.! How can an inorganic compound be use as an energy source. Respiration Review Electron Tower per cyto 2NADH 2H+ 2H+ low E o electron flow hi E o Q 2NAD+ H+ H+ O 2 4H+ H+ H+ 2 H 2 O ATP 3H+ ADP

2 Anaerobic 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 than oxygen 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 on Oxygen Concentrations! Points of reference: Atmospheric oxygen is ~21% (v/v) (or 2.1 x 10 5 parts per million) Low solubility in water: up to 14 parts per million (T and P dependent)! 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 human feces

3 Diversity of electron acceptors for respiration! Organic compounds: Eg. fumarate, dimethylsulfoxide (DMSO), Trimethylamine-Noxide (TMAO)! Inorganic compounds: Eg. NO 3-, NO 2-, SO 4 2-, S 0, SeO 4 2-, AsO 4 3-! Metals: Eg. Fe 3+, Mn 4+, Cr 6+! Minerals/solids: Eg. Fe(OH) 3, MnO 2! Gasses: Eg. NO, N 2 O, CO 2 Why is there so much diversity? How can prokaryotes accomplish this? Answer: Electron donor modules Electron acceptor modules Cyt b, Fe/S, FAD fumarate Fumarate reductase Dehydrogenase: Lactate Succinate Formate NADH Glycerophosphate Hydrogenase MQ UQ Cyt b, Fe/S, Mo DMSO reductase Cyt b, Fe/S, Mo DMSO TMAO TMAO reductase Cyt b, Fe/S, Mo NO 3 - Nitrate reductase

4 Modularity of electron transport chains what do most of these have in common? Example 1. Nitrate reduction

5 Figure Nitrogen cycle 78% N 2 Nitrate reducing bacteria! Contribute to denitrification (removal of?)! Beneficial process for sewage treatment plants Nitrogenous 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 Nitrogen gas: Pseudomonas stutzeri E V compared to for 1/2O 2 /H 2 O How many electrons are used from NO 3 - to N 2?

6 Dissimulative NO 3 - reduction Denitrification by Pseudomonas stutzeri

7 Denitrification by Pseudomonas stutzeri! Four terminal reductases Nap: Nitrate reductase (Mo-containing enzyme) Nir: Nitrite reductase Nor: Nitric oxide reductase N 2 or: Nitrous oxide reductase! All can function independently but they operate in unison Dissimilatory nitrate reduction: Biochemistry! Electron donor: lactate, formate, H 2, others Uses special dehydrogenases for these.! Enzymes are membrane-bound! Periplasmic nitrate reductases (NapA) contains a molybdenum cofactor! Coupled to the generation of PMF! ATP synthesized by oxidative phosphorylation

8 Nitrate vs. oxygen vs. denitrification respiration oxygen nitrite denitrification How much energy is made by reducing nitrate to nitrite with NADH? Nitrate (NO 3- ) example?e-?nad+ What s reduce and oxidized? Nitrite (NO 2- )?ATP?NADH Determining oxidation state of N and # of electrons: Nitrate= N(x) + 3O 2-! x + 3(-2) = -1! x= Nitrite= N(x) + 2O 2-! x + 2(-2) = -1! x= We only need to oxidize NADH for this: NADH + H + NAD + + 2H + + 2e- Find!Eo of nitrate/nitrite and NAD+/NADH Use Nernst Eq to find!go

9 Example 2. Arsenate reduction E o V Arsenate " arsenite Arsenic respiring bacteria and human health! Arsenic is mainly a groundwater pollutant! Affects ~140 million people among ~70 countries! Arsenate (As[V]): Like phosphate: H 2 AsO 4 - Affects ATP synthesis! Arsenite (As[III]): H 3 AsO 3 More toxic than As(V) Binds proteins Causes DNA damage! Microbes respire arsenate and make arsenite! Medical Geology problem

10 Respiring AsO 4 3- Shewanella sp. strain ANA-3 2As(V) Lactate Dividing cell 2As(III) As 2 S 3 Acetate + CO 2 Respiring O 2 Isolation of strain Example 3. Iron

11 Iron oxide reducing bacteria 2Fe(III) Fe III -oxide 2Fe(II) OM c-heme! Examples: Geobacter, Shewanella, Rhodoferrax! How do they do it? Q QH 2 CM NADH2 NAD+ Chemolithotrophy and Oxidation of Inorganic Molecules A. A pathway used by a small number of microorganisms called chemolithotrophs B. Produces a significant but low yield of ATP C. The electron acceptor is commonly O 2, some others include sulfate and nitrate D. The most common electron donors are hydrogen, reduced nitrogen compounds, reduced sulfur compounds, and ferrous iron (Fe 2+ )

12 Geological, biological, and anthropogenic sources of reduced inorganic compounds supporting chemolithotrophs Typical habitats of chemolithotrohs: Near the interface of oxic/anoxic conditions Energy yields from various inorganic electron donors:

13 Oxidation of sulfur-compounds! E.g.: Sulfur oxidizing Thiobacillii Thiobacillus thiooxidans Thiobacillus ferrooxidans Reactions H 2 S + 2 O 2 >>> SO 4 2-!G kj/mol + 2H HS- + 1/2O 2 + H+ >>> S 0 + H 2 O -209 S 0 + H 2 O O 2 >>> SO H S 2 O H 2 O + 2O 2 >>> 2 SO H+ -818! Produces sulfuric acid (H 2 SO 4 ) Acidification of soil Dissolution of minerals, e.g. CaCO 3 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 there is a mixture of reduced and oxidized chemicals.