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Nitrate Proton Symporter
Found on root cells where intake of nitrate and 2 protons are taken up at a time
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High affinity nitrate transporter
Michaelis-Menten Kinetics
Nitrate uptake at low concentrations (<1 nM)
v = Vmax[S]/(km + [S])
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Low affinity nitrate transporter
Nitrate uptake at high concentrations (>0.5 nM)
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Leucoplast
Organelle/plastid w/ storage and/or bio-synthetic transformation functions
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How can nitrate reductase cause plants harm?
Uptake of chlorate and NR turns it to chlorite which is toxic to plants.
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The Nitrogen Cycle
Atmospheric nitrogen (~78%) taken up by nitrogen fixing bacteria or lost in marine sediments.
Nitrates form and are taken up by plants.
Ammonium salts from feces or decay can turn to nitrates.
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CHL1 Tranporter
Dual affinity nitrate transporter. When phosphorylated, it is high affinity (for low conc.), and when not phosphorylated, it is low affinity.
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Nitrogen oxidation states
- NH3, NH4 = -3
- NO2, HNO2 = +3
- NO3, HNO3 = +5
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Nitrate in root cells
1. Short term storage in vacuoles.
2. Sent to the xylem.
3. Nitrate reduced by nitrate reductase to nitrite and taken up by leucoplast to turn to amides and sent through xylem.
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Nitrogen assimilation in leaves (mesophyll cells)
Taken in by 1 proton and nitrate symporter. Can be stored in vacuole.
Can be turned to nitrite by NR and sent to chloroplast to be turned to ammonium and be turned to amino acids.
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Nitrate Reductase
NO3 -> NO2 + H2O
(N-terminus)
- MoCo (molybdenum complex)
- Heme~Cytb557
- FAD
(C terminus)
NADH + H --> NAD(+) + 2e (H)
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Leucoplast nitrite reaction
NO2 -[Nitrite reductase]-> NH4 --> Amide
3NADPH + 3H required during nitrite reductase
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Chloroplast nitrite reaction
NO2 -[6 Ferrodoxin; Nitrite reductase]->NH4 --> AA
Ferrodoxin oxidized
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Regulation of nitrate reductase
Gene regulation
Enzyme Regulation
- Gene regulation
- +: Light, glucose (or other carbs), nitrate
- -: Glutamine, NH4, other AA
- Enzyme Regulation (phosphorylation leads to inhibition)
- +: Stimuli~Ca, okadaic acid
- -: Triose phosphate, phosphate ester
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Nitrite Reductase
Monomeric enzyme w/ 2 redox centers
(N terminus)
Fdx(red) --> Fdx(ox)
Fdx-binding domain (redox center 1)
4Fe-4S, Siroheme (redox center 2)
(C terminus)
NO2 --> NH4
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Coupling of photosynthetic e to nitrite reduction in chloroplast
Light oxidized fdx which is needed for nitrite reductase.
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Oxidative pentose phosphate pathway role in leucoplast
3 NADPH comes from PPP which is used to oxidize fdx
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Glutamine Synthetase-Glutamine 2-oxoglutarate amino transferase (GS-GOGAT) pathway
NH4 + glutamate -[glutamine synthetase]->
Glutamine + 2-oxoglutarate -[glutamate synthase]->
2 Glutamate
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Glutamine synthetase requirements
Glutamate synthase requirements
ATP
- NADPH in nonphotosynthetic tissues
- Fdx(red) in chloroplast
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Liberty links crops
Herbicide tolerant for weed control ~ soybean, corn, cotton, canola
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Glutamate Dehydrogenase Pathway
NH3 + 2-oxoglutarate <-[Glutamate dehydrogenase]-> Glutamate + H2O
Forward reaction requires NAD(P)H
Mainly used in deaminating glutamate (reverse) for nitrogen reallocation
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Amino groups synthesized from transamination reactions
Glutamate + oxaloacetate -[Aspartate aminotransferase]-> Aspartate + 2-oxoglutarate
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Nitrogen fixing organisms and host plant
- Legumes: Rhizobium
- Sugar cane: Acetobacter
- Azolla: Anabaena
Cyanobacteria
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Nitrogenase enzyme complex
Fe protein and MoFe protein: Dinitrogenase reductase and dinitrogenase
16 ATP --> 2NH3, H2 from N2 + 8H
Glutamine and Aspargine are main products
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Crop rotation
legunes -> roots -> fruits -> leaves
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Nitrogen storage forms
- Vacuole (short)
- Aspargine + homoserine (nonprotein AA) (med)
- Storage protein (patatin) (long)
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