17' Nitrogen fixation and amino acid biosynthesis

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17. Nitrogen fixation and amino acid biosynthesis
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Nitrogen cycles between oxidized reduced forms
in the biosphere
degradation (animals microorganisms)
synthesis (microorganisms, plants animals)
(Rhizobium some other bacteria)
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The roots of leguminous plants have nodules
containing N2-fixing bacteria
Lehninger Fig. 22-4a
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Bacteroids containing nitrogenase are found
inside the nodule cells
bacteroids (rod-like bacteria)
plant cell nucleus
Lehninger Fig. 22-4b. (This electron micrograph
is colorized artificially.)
Nitrogenase is very sensitive to O2. It is
protected in the nodules by a high concentration
of leghemoglobin, a heme protein with a strong
affinity for O2. Leghemoglobin is produced by
the plant, but carries electrons to the bacterial
respiratory chain, keeping the O2 concentration
low.
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Nitrogenase from Azobacter vinelandii contains
iron-sulfur and iron-molybdenum centers
Azotobacter are free-living, aerobic soil
bacteria.
Fe-Mo protein FeS cluster and Fe-Mo cofactor
Fe protein FeS cluster two ATP-binding sites
structurally homologous to G-proteins
1n2c.pdb
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Nitrogenase from Azobacter vinelandii
Mo-Fe-S cluster (Mo7Fe9S)
8Fe7S cluster
4Fe4S cluster
Mg ADP (2)
1n2c.pdb
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The Fe-Mo cofactor
Cys residue of the protein
Intermediates in which partially reduced
derivatives of N2 replace one of the O atoms
bound to the Mo have been proposed, but the
mechanism of the N2-fixation reaction is not
known.
Homocitrate (3-hydroxy-3-carboxyadipic acid)
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Nitrogenase uses 8 electrons and 16 ATP to
reduce N2 2 H to 2 NH4 H2
The Fe protein transfers one electron at a time
to the Fe-Mo protein.
The ATP stoichiometry is uncertain. Only 8 ATP
are needed under some conditions.
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Ammonia is incorporated into many biological
molecules through glutamine and glutamate
(2)
H2O
Glu
(3)
a-keto-glutarate
(1)
a-keto-glutarate
Glutamate dehydrogenase (1) and glutamine
synthetase (2) are found in all organisms.
Reaction (3) occurs in plants bacteria, but not
animals.
Glu
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Glutamine synthetase catalyzes formation of
glutamine from glutamate and NH4
NH4 Pi
ATP ADP
Glu
Gln
The reaction proceeds through an enzyme-bound
?-glutamylphosphate intermediate
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Glutamine serves as a donor of amine groups for
synthesis of many other molecules
In most terrestrial animals, Gln also carries
ammonia in the blood to the liver kidneys for
excretion as urea.
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Glutamine synthetase is inhibited allosterically
by many of the end-products
carbamoyl-phosphate
Gln
Glu
glutamine synthetase
glucosamine-6-P
ATP ADP NH4 Pi
alanine
glycine
histidine
tryptophan
The inhibitory effect of all the products acting
in concert is greater than the sum of their
individual effects.
AMP
CTP
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E. coli glutamine synthetase also is controlled
by covalent modification
Gln
adenylylation
deadenylylation
a-keto-glutarate
adenyl group
The regulation by Gln and a-ketoglutarate
involves similar covalent modifications
(uridylylation) of the enzymes that add or remove
the adenyl group.
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Bacterial glutamine synthetase has 12 identical
subunits
views of the Salmonella typhimurium enzyme
parallel and perpendicular to the 6-fold symmetry
axis
2gls.pdb
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Humans can synthesize 10 of the 20 common amino
acids
Arg is essential in infants and growing children
but not in adults.
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Building blocks for amino acid synthesis in
humans come from glycolysis and the citric acid
cycle
glucose
glycine
3-phosphoglycerate
serine
cysteine
pyruvate
alanine
proline
citrate
oxaloacetate
glutamate
glutamine
a-ketoglutarate
aspartate
arginine
asparagine
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Serine is formed via 3-phosphoglycerate
3-phosphoglycerate
3-phosphohydroxypyruvate
NADH
NAD
Glu
a-kG
serine
3-phosphoserine
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Plants and bacteria synthesize aromatic amino
acids from carbohydrates via shikimic acid