Flashcards in Nitrogen Metabolism Deck (29):
Nitrogen in the atmosphere
- most abundant element in air (80%)
- occurs as dinitrogen
- convert N2 to ammonia
- conversion of N2 into NH3-
- the "fixed" nitrogen is immediately incorporated into organic compounds
- only prokaryotes fix nitrogen
- NO nitrogen fixing plants or animals.
- release of ammonia into the environment from N-containing organic compounds
- oxidation of ammonia by aerobic bacteria
- reduction of nitrate to N2 by anaerobic respiration
biological Nitrogen fixation reaction
- N2 + 8H+ + 8e- + 16-32 ATP -> 2 NH3+ H2 + 16-32 (ADP + Pi)
Non biological nitrogen fixation
- N2 + 3H2 -> 2 NH3
- Haber-Bosch reaction
- accounts for 1/5 of nitrogen fixed per year
Why is nitrogen fixation so expensive?
- N2 is joined by a very stable triple bond that is difficult to break.
Who fixes nitrogen?
- Free living heterotrophs - Azotobacter and Pseudomonas
- Phototrophs - Anabaena, Nostoc, Rhodospirrilum, Rhodobacter
- Plant symbiotic bacteria
- Rhizobium - leguminous plant
- Frankia - angiosperms
- Azospirillum - grass, maize
- carries out nitrogen fixation
nitrogen is reduced at
- molybdenum-iron cofactor
- reducing power stored in P-cluster - stores electrons
- you must make 26 proteins to support N2 fixation
- protein synthesis is expensive
- it takes 16-32 ATP + NADH per N2 fixed
- regulated at transcriptional level and post-translational level
- NtrB and NtrC two component regulatory system
- NtrB is the sensor - senses N status of the cell
- NtrC is the response regulate - activates transcription of NifA, NifL
- NifA - positive regulator
- NifL - negative regulator
nitrogen status measured by
- measured to the ratio of alpha-ketoglutarate/glutamate via PII
Transcriptional regulation process
- nitrogenase extremely sensitive to O2. Only turn on if O2 is low
- in presence of high levels of O2, NifL will bind to NifA and not allow it to turn on transcription of nitrogenase operon
- If O2 levels are low, NifL will not bind NifA, and it can activate transcription
Post translational regulation
- the cell shuts off nitrogenase whenever ammonia feels increase by modifying an arginine residue in the active site of the iron protein.
- done by adding an ADP ribose via DRAT.
- when ammonia levels drop, the ADP ribose removed by DRAG.
Nitrogenase and O2
- nitrogenase is rapidly inactivated by exposure to O2
- The FeMoCo center binds to O2 tightly, and the entire enzyme must be degraded and made again
- Many N2 fixers have developed mechanisms to protect nitrogenase from O2
- obligate aerobe, but can fix N2
- to protect nitrogenase from inactivation use
- respiratory protection
- conformational protection
- Azotobacter keeps its cytoplasm at very low O2 concentration through its ETC
- At high O2 levels, it will turn on an alternative terminal oxidase, cytochrome bd, that has a high affinity for O2 and a very rapid rate of O2 reduction
- but it's not a proton pump, so less energy available
- if O2 is too high, FeSII will bind to nitrogenase and protect the FeMO cluster from O2 inactivation
- When FeSII is bound to the protein, N2 also does not have access and nitrogen fixation stops
- When O2 drops, FeSII dissociates and nitrogen fixation proceeds.
- cannot fix nitrogen and photosynthesize at the same time because O2 will damage nitrogenase
- when these bacteria need to fix nitrogen, they do so in a heterocyst and share nitrogen with neighbors who share their fixed CO2
- no photosynthesis occurs in a heterocyst so nitrogenase is not inhibited.
- nitrogen fixing bacteria live in symbiosis with plants
- environment inside the nodule kept at low O2 by leghemoglobin
lock and key mechanism
- only bacteria that secrete the correct polysaccharide "key" are able to colonize each host.
- plant secretes flavonoids that stimulate bacteria to start producing Nod factors
- bacteria make polysaccharide which induces root hair curling and allows the bacteria to invade into the cortex cells.
- will invade cell and divide rapidly producing a nodule on the root.
- once N2 is fixed to ammonia, it is almost immediately incorporated into amino acids
- when an organism is growing on high nitrogen:carbon ratio diet
- will excrete ammonia
- aerobic process
- bacteria of the genus nitrosomas and nitrobacter oxidize ammonia to nitrite and nitrate
- gets energy from oxidation of ammonia to nitrite
- does in two steps
- first step produces hydroxylamine
- then oxidizes to nitrite
- uses nitrite as energy source to produce nitrate