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N2 + 3H2 ---> 2NH3 ∆G’º = -33.5 kJ/mol
However, N-N triple bond is a significant kinetic barrier
Bond energy = 930 kJ/mol
N2 + 10 H+ + 8 e- + 16 ATP ---> 2 NH4+ + 16 ADP + 16 Pi + H2
Nitrogen is fixed by anaerobic bacteria
0 -3
Proposed mechanism for dinitrogen reduction
Mo
FeS
N
N
Mo
N
N
FeS
H+
Mo
N
NH
FeS
H+
Mo
N
NH2
FeS
+ H+
Mo
N
NH3+
FeS
NH3
Mo
N
FeS
Mo
N+
Mo
N+
Mo
N+
Mo
N+
Mo
N+
H+
Mo
NH
FeS
H+
Mo
NH2
FeS
H+
Mo
NH3
FeS
Nitrate:ferredoxin reductase
S
Mo4+ O
O-
N+
O
O-
S
Mo6+ O
O-
N
O
O-
S
Mo6+ O
O2-
S
Mo6+ O
OH-
H+
H-
S
Mo4+O
H2O
Dissimilatory/Respiratory Nitrite Reductase
Siroheme and ferredoxin dependent looks and behaves a lot
like cytochrome c oxidase
NO2- ----> NO ----> NH2OH ----> NH4
+
Allows anaerobic organisms to respire electrons onto nitrite instead of oxygen
+3 +1 -1 -3
Dissimilatory Nitrite Reductase
NO2- ----> NO
Produces NO and N2O - potent greenhouse gasses
+3 +2
NO ---> N2O+2 +1
Nitrite reductase, multicopper enzyme
Nitric oxide reductase, heme dependent
N2O ---> N2
+1 0
Nitrous oxide reductase, multicopper enzyme
Biological Nitrification
NH3 ---> NO2- ---> NO3
-
-3 +3 +5
NH3 + O2 + 2 e- + 2 H+ ---> NH2OH + H2O
Ammonia monooxygenase (related to methane monooxygenase)
NO2- ---> NO3
- is non-enzymatic
NH2OH + H2O ---> NO2- + 5 H+ + 4 e-
Hydroxylamine oxidoreductase (heme dependent)
2 of these electrons are transferred to cytochrome bc1
Ultimately they land on O2 and generate proton gradient
2 electrons are returned to AMO to complete the cycle
Glutamine synthase
glutamate + NH4+ + ATP ---> glutamine + ADP + Pi + H+
Glutamate synthase
glutamine + KG + NADPH + H+ ---> 2glutamate + NADP+
Net reaction
KG + NADPH + NH4+ + ATP ---> glutamate + NADP+ + H2O + ADP + Pi
Need catalytic amounts of glutamate!There has to be another way!
H
COO-
CH2
CH2
NH3+
O O-
ATP
H
COO-
CH2
CH2
NH3+
O OPO32-
H
NH2
BH
COO-
CH2
CH2
NH3+
O NH2
Glutamine synthase
Glutamate synthaseCOO-
C+H3N H
CH2
CH2
C
NH2O
E
S
HB
H A
COO-
C+H3N H
CH2
CH2
C
OSE
+NH3
H2O
COO-
C+H3N H
CH2
CH2
C
O-O
COO-
CO
CH2
CH2
C
O-O
COO-
C+H2N
CH2
CH2
C
O-O
H-HA
COO-
C+H3N H
CH2
CH2
C
O-O
What about the reverse reaction of glutamate dehydrogenase?
glutamate + NADP+ + H2O ---> KG + NH4+ + NADPH
Normally this reaction favors the right
Glutamine synthetase is a control point for growth. Many end products allosterically regulate its activity
N
NN
N
NH2
O
OHOH
HH
HH
OP-O
O
O
P-O O
O
P O
O-
-OC
O
O-HO
N
NN
N
NH2
O
OHOH
HH
HH
OP-O
O
O
P-O O
O-
C
O
OHO P
O
O-
O-
Carbamoylphosphate synthase: step 1
Carbonic phosphoric anhydride
Bicarbonate
Carbamoylphosphate synthase: step 2
N
NN
N
NH2
O
OHOH
HH
HH
OP-O
O
O
P-O O
O
P O
O-
-O
C
O
OHO P
O
O-
O-
HN
H
H
H2N
C
O
O
H
BN
NN
N
NH2
O
OHOH
HH
HH
OP-O
O
O
P-O O
O-H2N
C
O
O P
O
O-
O-
Carbonic phosphoric anhydride
Carbamoylphosphate
Carbamate
Ornithine transcarbamoylase
+ Pi
OrnithineCitrulline
These are -amino acids
N+ CH2 CH2 CH2 CH
NH3+
COO-
H2N C
O
O P
O
O-
O-
H
H
HB
HN CH2 CH2 CH2 C
H
NH3+
COO-C
O
H2N
Argininosuccinate synthase: step 1
N
NN
N
NH2
O
OHOH
HH
HH
OP-O
O
O
P-O O
O
P O
O-
-O
N
NN
N
NH2
O
OHOH
HH
HH
OPO
OH
O
H C
COO-
NH3+
CH2
CH2
CH2
NH
C
NH
H
B
O-
P-O O
O
P O
O-
-O
NH2+
C
NH
CH2
CH2
CH2
CH NH3+
COO-
O
Citrulline Citrullyl-AMP
Argininosuccinate synthase: step 2
Citrullyl-AMP
N C
COO-
H
CH2
COO-
H
H
H
B
B
N
NN
N
NH2
O
OHOH
HH
HH
OPO
OH
ONH2+
C
NH
CH2
CH2
CH2
CH NH3+
COO-
N
NN
N
NH2
O
OHOH
HH
HH
OP-O
OH
O
NH2+
C
NH
CH2
CH2
CH2
CH NH3+
COO-
HN C
COO-
CH2
COO-
H
Argininosuccinate
NH2+
C
NH
CH2
CH2
CH2
CH NH3+
COO-
HN C
COO-
C
COO-
H
HH
B
A
H
NH2+
C
NH
CH2
CH2
CH2
CH NH3+
COO-
NH2
C
C
-OOC H
H COO-
Argininosuccinate lyase
Argininosuccinate Arginine
Fumarate
NH2+
C
NH
CH2
CH2
CH2
CH NH3+
COO-
NH2H
OH
B NH2
C
NH
CH2
CH2
CH2
CH NH3+
COO-
NH2O
H
B
H
A
NH2
C
NH2
CH2
CH2
CH2
CH NH3+
COO-
NH2
O
Arginase
In respiration asp/mal shuttle
Moves NADH into the mitochondrionwith net movement of one H+ in
but also moves NH3 out!
For every asp out a mal goes inand NH3 is moved out to cytoplasm
S2- + cyt c553(ox) ---> S + cyt c553(red)
Sulfide dehydrogenase
Sulfide:quinone oxidoreductase
S2- + UQ(ox) ---> S + UQH2(red)
S2- ---> S ---> SO42-
In photosynthetic green sulfur bacteria this oxidation is the driving force for the rTCA cycle
Elemental sulfur globules are temporarily deposited outside the cell.
Sulfur oxidation
Sulfur oxygenase reductase (SOR)
Mononuclear non-heme iron enzyme
SORS + O2 ----> HSO3
- + H+
Non-enzymatic disproportionation3S + 3H2O ----> HSO3
- + 2HS- + 3H+
Sum4S + O2 + 4H2O ----> 2HSO3
- + 2HS- + 4H+
Non-enzymatic reaction with more SS + HSO3
- ----> S2O32- + H+
Sulfite oxidase converts sulfite to sulfate
S
Mo6+S
O2-
O
O
S O-
O-
S
Mo4+S
O-
O
O
S+ O-
O-
S
Mo4+S
-O
O
O
S O-
OO
H2O
Fe3+
S
Mo5+S O
-OH
cytochrome c
S
Mo5+S O
-OH
Fe3+S
Mo6+S O
O-H
HH
B
B
Thiosulfate:quinone oxidoreductase
Tetrathionate
S4O62- + H2O ---> S3O3
2- + SO42- + 2H+
Tetrathionate hydrolase
S3O32- ---> S2O3
2- + S
Spontaneous disproportionation
2S2O32- S4O6
2-
E S
H
S
H
-O S
O
O
O P
O
O-
A
B
E S
S
H
S
O
O
O-
B
E S
S
S
O
O-
O-
H-
H+
E S
H
S
H
NADPH reduces the disulfide
PAPS Reductase
Sulfate/ite respiring organisms use dissimilatory sulfate/ite reductase
This is a respiratory enzyme that couples sulfate/ite reduction with NADPH
oxidation via ferredoxinrelated to cytochrome c oxidase
Catalyzes the reduction of sulfite to sulfide
NH+
O-
2-O3PO
CH O
H C
COO-
NH3+
C
O
H
H
Schiff BaseFormation
NH+
O-
2-O3PO
CH
H C
COO-
NH+
C
O
H
HB
HA
NH+
O-
2-O3PO
CH
C
COO-
NH+
C H
H
+ acetate
Serine
O
CH3
O
CH3
HS-
NH+
O-
2-O3PO
CH
C
COO-
NH+
C H
H
SHH+
H
Acetylserine lyase
+H3NC
COO-
H
S
CH3
+H3NC
COO-
H
C O-
O
+H3NC
COO-
H
C H
O
+H3NC
COO-
H
CH2
HO
+H3NC
COO-
H
CH2
OC
O
H2C
CH2
COO-
Aspartate Aspartate-γ-semialdehyde
HomoserineO-succinylhomoserine
succinyl-CoA
+H3NC
COO-
H
CH2
SCH2
CCOO-+H3N
H
Cystathionine
cysteine
succinate
Cystathionineγ-synthase
+H3NC
COO-
H
CH2
HS
Homocysteine
Cystathionineβ-lyase
Pyruvate + NH3
S-adenosylmethionine
S-adenosylhomocysteine
serine
Cystathionineβ-synthase
cysteine+
-ketobutyrate
Cystathionineγ-lyase
Methionine
Cystathionine-γ-synthaseCOO-
H
NH+
H2C O C
H2C
H2C COO-
H
NH+
B
COO-
NH+
H2C O C
H2C
H2C COO-
H
NH
H2C
H2C O
O
H2C
H2C COO-
O
O
H
COO-
NH3+
H2C
H2C
H+
COO-
NH+
H2C O C
H2C
H2C COO-
HC
H
NH+
O
H2C
Cystathionine-γ-synthase
COO-
NH+
H2C O C
H2C
H2C COO-
HC
H
NH+
O
CH
H
B
COO-
NH+
CH2
-O CH2C
H2C COO-
HC
H
NH+
O
CH
H SH2C
COO-
H
NH3+B
COO-
NH
H2C
HC
H
NH+
CH
SH2C
COO-
H
NH3+
H+COO-
NH+
H2C
HC
H
NH+
H2C S
H2C
COO-
H
NH3+
Cystathionine-γ-synthase
COO-
NH+
H2C
HC
H
NH+
H2C S
H2C
COO-
H
NH3+
BH+
COO-
NH+
H2C
HC
NH+
H2C S
H2C
COO-
H
NH3+
H
Cystathionine-β-lyase
COO-
H
NH3+
H2C S
H2C
H2C
COO-
H
NH3+
COO-
H
NH+
H2C S
H2C
H2C
COO-
H
NH3+H
NH+
B
COO-
NH+
H2C S
H2C
H2C
COO-
H
NH3+H
NH
H+COO-
NH+
CH2 HSH2C
H2C
COO-
H
NH3+H
NH+
HN
CH2
COO-
H
B
H+
H3C
NH2+
COO-
Homocysteine
N
HN
HNCH3
Co+NN
N N
HA
Co3+NN
N N
CH3
Co3+NN
N N
CH3
+H3NC
COO-
H
S
H
B
Co+NN
N N
+H3NC
COO-
H
S
CH3
Methionine synthase
In mammals cysteine is made from methionine
+H3NC
COO-
H
S
CH3
+H3NC
COO-
H
C O-
O
+H3NC
COO-
H
C H
O
+H3NC
COO-
H
CH2
HO
+H3NC
COO-
H
CH2
OC
O
H2C
CH2
COO-
Aspartate Aspartate-γ-semialdehyde
HomoserineO-succinylhomoserine
succinyl-CoA
+H3NC
COO-
H
CH2
SCH2
CCOO-+H3N
H
Cystathionine
cysteine
succinate
Cystathionineγ-synthase
+H3NC
COO-
H
CH2
HS
Homocysteine
Cystathionineβ-lyase
Pyruvate + NH3
S-adenosylmethionine
S-adenosylhomocysteine
serine
Cystathionineβ-synthase
cysteine+
-ketobutyrate
Cystathionineγ-lyase
Methionine
H
COO-
NH3+
H2C S+
OA
OH OH
H2C
CH3
Nu-
H
COO-
NH3+
H2C S
OA
OH OH
H2C
Nu-CH3H
O H B
H+
H
COO-
NH3+
H2C SH
OA
OH OH
H2C HO
SAM-methyltransferase/S-adenosylhomocysteine hydrolase
HOH2C
COO-
H
NH3+
H2C
COO-
H
NH+H
NH+
B
HOH2C
COO-
NH+H
NH
H2C
COO-
NH+
NH+
HO
H+
COO-
NH3+
HH2C
H2C S
H
B
H+
COO-
H
NH3+
H2C S
H2C
H2C
COO-
H
NH+H
NH+
Cystathionine-β-synthase
COO-
H
NH3+
H2C S
H2C
H2C
COO-
H
NH3+
COO-
H
NH3+
H2C S
H2C
H2C
COO-
H
NH+H
NH+
B
COO-
H
NH3+
H2C S
H2C
H2C
COO-
NH+H
NH
H+
COO-
H
NH3+
H2C S
H2C
H2C
COO-
NH+
NH+
Cystathionine-γ-lyase
Cystathionine-γ-lyase
COO-
H
NH3+
H2C S
H2C C
H
COO-
NH+
NH+
H
B
COO-
H
NH3+
H2C S
H2C C
H
COO-
NH
NH+
H+
COO-
H
NH3+
H2C SH H2C C
H
COO-
N
NH+
HB
H3C CH
COO-
N
NH+
H+
COO-
NH2+
H
B
H+
COO-
NH2+
COO-
O