11576442
Description
| Question | Answer |
| Limiting nutrients for plant growth and productivity | Nitrogen, water, carbon |
| Considering nitrogen is so abundant in the atmosphere, why is it a limiting factor in plant growth? | N2 is very stable but must be fixed/reduced to be used by plants |
| basic reaction of Nitrogen fixation | N2 reduced to NH4+ (NH3), which can then be converted to biomolecules |
| Haber Bosch industrial process | Nitrogen and hydrogen reacted over iron at 350C and >200atm N2(g) + 3H2(g) ↔ 2NH3(g) fertilizer from this supports 1/3 of our population! |
| Steps in nitrogen cycle performed exclusively by bacteria and archaea | Nitrogen fixation: N2 --> NH3 Nitrification: NH3 --> NO3- |
| Steps in nitrogen cycle that are not necessarily performed by bacteria or archaea | Denitrification: NO3- --> N2 Ammonification/Assimilation & Assimilation: NH3 --> NH2 groups, organic compounds and proteins --> NO3- |
| chemical fertilizer PROS | INCREASED CROP YIELD |
| chemical fertilizer CONS | fertilizer CONTAMINATES ground water and consumes fossil fuel resources!! |
| Chemical vs Biological N fixation | N2 triple bond is hard to break, even though ΔG is negative a lot of energy (high Temp, Pressure, and iron catalyst) is required for reaction biological fixation occurs at low Temp, 1 atm, with a nitrogenase enzyme as the catalyst; energy supplied from ATP |
| Equation for biological nitrogen fixation | N2 + 8H+ + 8e- + 16MgATP --> 2NH3 + 16MgADP + 16Pi + H2 e- come from reduced Fd or flavodoxin these numbers are all a minimum!! 16-24 ATP typically used |
| Options for nitrogenase reduction reaction | Nitrogenase can reduce N2 to ammonia or acetylene to ethylene: 1) N2 + 8H+ + 8e- --> 2NH3 + H2 2) C2H2 + 2H+ + 2e- --> C2H4 *both break a triple bond* |
| How to demonstrate nitrogenase activity | digest cells containing stable isotope 15N2 --> they will release NH3 from biomolecules --> shown to contain 15N by mass spec OR (easier) reduce acetylene to ethylene with analysis in a gas chromatograph shows production of C2H4 from C2H2 over time |
| How do you get DEFINITIVE proof of N2 fixation | grow cells in sealed containers, give air at top if aerobic or regular N2 gas/etc for anaerobic → inject nitrogenase → after periods of time withdraw samples from gas and see if acetylene (C2H2) has been converted to ethylene (C2H4) |
| proteins within nitrogenase complex | dinitrogenase reductase = Fe protein, accepts e- from Fdred and binds ATP --> passes e- to FeMo and hydrolyzes ATP dinitrogenase = FeMo protein, uses e- to reduce bound N2 to NH3 *both rxn centers VERY susceptible to O2* |
| How many ATPs are hydrolyzed when e- move from Fe to FeMo cofactor? | 4 ATP hydrolyzed per step *4 steps total, so 24 ATP used up* |
| Nitrogenase complex reaction cycle | 1) Fe protein gets 2e- from an e-donor (Fdox) and binds 4 ATP 2) Transfers 2 e- to FeMo protein and hydrolyzes 4 ATP 3) FeMo protein binds 2H+ --> reduced by the e- to H2 4) N2 displaces H2 on the FeMo protein 5) Fe protein reduced by Fd(red) 3X more times, total of 8e- and 16 ATP 6) at each turn of 5, 2e- transferred to bound N2, yielding 2NH3 7) NH3 is found as the soluble ion NH4+ |
| How do phototrophic organisms reduce ferrodoxin? (considering Fdred is a strong donor) | Cyanobacteria use light energy and PSI to create Fd(red) *review end of Z-scheme* |
| How do non-phototrophic ANAEROBES organisms reduce ferrodoxin? (considering Fdred is a strong donor) | use pyruvate-ferrodoxin oxidoreductase Pyruvate + CoA + Fd(ox) → Acetyl-CoA + CO2 + Fd(red) |
| How do non-phototrophic AEROBES organisms reduce ferrodoxin? (considering Fdred is a strong donor) | use reverse electron transfer from NADH/anything with higher reduction potential than Fd Energy in the form of ATP or PMF is used! --> may account for the fact that nitrogen fixation uses more than the predicted 16 ATP per 2NH3 |
| How do cyanobacteria protect nitrogenase from O2, considering their phototrophy produces O2? | 1) Temporal separation of photosynthesis and nitrogen fixation 2) Spatial separation of the two processes: vegetative cells for photosynthesis and heterocysts for nitrogen fixation |
| heterocysts | terminally differentiated cell in cyanobacteria where photosystem II is dismantled and oxidases are produced to keep the O2 concentration very low |
| How do obligate aerobes protect nitrogenase from O2? | A. vinelandii : obligate aerobe soil bacteria 1) produces slime layer --> limits O2 influx in cell 2) VERY high respiratory rate --> removes O2 quickly 3) protective protein binds to nitrogenase and protects it from damage by O2 *most important of the three* |
| How do symbionts protect nitrogenase from O2? | S. meliloti: aerobic gram negative symbiont of alfalfa grass --> form root nodules that develop into bacteroids with N2-fixing ability --> in nodule, leghemoglobin produced by the plant binds O2 and reduces free [O2] *free-living S. meliloti does not have genes required for N2 fixation in presence of O2* |
| nif regulon | genes required for N2 fixation! ONLY expressed when needed (no NH4+) & when they could function properly (low O2) NifA is positive regulator; control of expression (via NtrC) and activity (via NifL) important for appropriate response to environmental conditions |
| N2 fixation regulation in FREE-LIVING Klebsiella pneumoniae | Presence of O2 OR NH4+ blocks nif tx!! NtrB: His kinase activated by LOW [NH4+] NtrC: DNA-binding response regulator σ54: positive regulator of N-scavenging & N2-fixing genes, works with NtrC NifA: positive regulator of nif genes NifL: negative regulator of NifA, binds to NifA in presence of O2 to block nif tx |
| how is NtrB kinase activity promoted? | cells are effectively looking at their C/N ratio *REVIEW THIS* |
| Regulation of nitrogen fixation genes in S. meliloti during symbiosis | O2 blocks FixL --> blocks nif and fix tx FixL: His kinase with heme sensing domain FixJ: DNA-binding response regulator NifA and FixK: tx factors activate expression of nif and fix genes Other nif and fix genes also involved! |
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