Lecture 8

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Lecture 8
2013-02-18 03:55:43
MCB 126

Nitrogen and sulfur assimilation and amino acid synthesis
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  1. Nitrate Proton Symporter
    Found on root cells where intake of nitrate and 2 protons are taken up at a time
  2. High affinity nitrate transporter

    Michaelis-Menten Kinetics
    Nitrate uptake at low concentrations (<1 nM)

    v = Vmax[S]/(km + [S])
  3. Low affinity nitrate transporter
    Nitrate uptake at high concentrations (>0.5 nM)
  4. Leucoplast
    Organelle/plastid w/ storage and/or bio-synthetic transformation functions
  5. How can nitrate reductase cause plants harm?
    Uptake of chlorate and NR turns it to chlorite which is toxic to plants.
  6. 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.
  7. CHL1 Tranporter
    Dual affinity nitrate transporter. When phosphorylated, it is high affinity (for low conc.), and when not phosphorylated, it is low affinity.
  8. Nitrogen oxidation states
    • NH3, NH4   = -3
    • NO2, HNO2 = +3
    • NO3, HNO3 = +5
  9. 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.
  10. 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.
  11. Nitrate Reductase
    NO3 -> NO2 + H2O


    • MoCo (molybdenum complex)
    • Heme~Cytb557
    • FAD

    (C terminus)

    NADH + H --> NAD(+) + 2e (H)
  12. Leucoplast nitrite reaction
    NO2 -[Nitrite reductase]-> NH4 --> Amide

    3NADPH + 3H required during nitrite reductase
  13. Chloroplast nitrite reaction
    NO2 -[6 Ferrodoxin; Nitrite reductase]->NH4 --> AA

    Ferrodoxin oxidized
  14. 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
  15. 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
  16. Coupling of photosynthetic e to nitrite reduction in chloroplast
    Light oxidized fdx which is needed for nitrite reductase.
  17. Oxidative pentose phosphate pathway role in leucoplast
    3 NADPH comes from PPP which is used to oxidize fdx
  18. Glutamine Synthetase-Glutamine 2-oxoglutarate amino transferase (GS-GOGAT) pathway
    NH4 + glutamate -[glutamine synthetase]->

    Glutamine + 2-oxoglutarate  -[glutamate synthase]->

    2 Glutamate
  19. Glutamine synthetase requirements

    Glutamate synthase requirements

    • NADPH in nonphotosynthetic tissues
    • Fdx(red) in chloroplast
  20. Liberty links crops
    Herbicide tolerant for weed control ~ soybean, corn, cotton, canola
  21. 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
  22. Amino groups synthesized from transamination reactions
    Glutamate + oxaloacetate -[Aspartate aminotransferase]-> Aspartate + 2-oxoglutarate
  23. Nitrogen fixing organisms and host plant
    • Legumes: Rhizobium
    • Sugar cane: Acetobacter
    • Azolla: Anabaena

  24. 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
  25. Crop rotation
    legunes -> roots -> fruits -> leaves
  26. Nitrogen storage forms
    • Vacuole (short)
    • Aspargine + homoserine (nonprotein AA) (med)
    • Storage protein (patatin) (long)