FCH 532 Lecture 26
Chapter 26: Essential amino acids
Quiz Monday: Translation factors
Quiz Wed: NIH Shift
Quiz Fri: Essential amino acids
Exam 3: Next Monday
Amino acid biosynthesis
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Essential amino acids - amino acids that can only be
synthesized in plants and microorganisms.
Nonessential amino acids - amino acids that can be
synthesized in mammals from common intermediates.
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Table 26-2
Essential and Nonessential Amino Acids in
Humans.
Nonessential amino acid
biosynthesis
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Except for Tyr, pathways are simple
Derived from pyruvate, oxaloacetate, -ketoglutarate, and 3phosphoglycerate.
Tyrosine is misclassified as nonessential since it is derived
from the essential amino acid, Phe.
Glutamate biosynthesis
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Glu synthesized by Glutamate synthase.
Occurs only in microorganisms, plants, and lower animals.
Converts -ketoglutarate and ammonia from glutamine to
glutamate.
Reductive amination requires electrons from either NADPH or
ferredoxin (organism dependent).
NADPH-dependent glutamine synthase from Azospirillum
brasilense is the best characterized enzyme.
Heterotetramer (22) with FAD, 2[4Fe-4S] clusters on the 
subunit and FMN and [3Fe-4S] cluster on the subunit
NADPH + H+ + glutamine + -ketoglutarate  2 glutamate + NADP+
Figure 26-51 The sequence of reactions catalyzed by
glutamate synthase.
Electrons are
transferred from
NADPH to FAD at
active site 1 on the 
subunit to yield FADH2.
2.
Electrons transferred
from FADH2 to FMN on
site 2 to yield FMNH2.
3.
Gln is hydrolyzed to glutamate and
ammonia on site 3 of
the  subunit.
4.
Ammonia is
transferred to site 2 to
form -iminoglutarate
from -KG
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1.
5.
-iminoglutarate is
reduced by FMNH2 to
form glutamate.
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Figure 26-52 X-Ray structure of the  subunit of A.
brasilense glutamate synthase as represented by its C
backbone.
Figure 26-53 The  helix of A. brasilense glutamate
synthase.
C-terminal domain of
glutamate synthase is a 7turn, right-handed  helix.
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43 angstrom long.
Structural role for the
passage of ammonia.
Ala, Asn, Asp, Glu, and Gln are
synthesized from pyruvate,
oxaloacetate, and -ketoglutarate
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Pyruvate is the precursor to Ala
Oxaloacetate is the precursor to Asp
-ketoglutarate is the precursor to Glu
Asn and Gln are synthesized from Asp and Glu by amidation.
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Figure 26-54 The
syntheses of alanine,
aspartate, glutamate,
asparagine, and
glutamine.
Gln and Asn synthetases
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Glutamine synthetase catalyzes the formation of glutamine
in an ATP dependent manner (ATP to ADP + Pi).
Makes glutamylphosphate intermediate.
NH4+ is the amino group donor.
Asparagine synthetase uses glutamine as the amino donor.
Hydrolyzes ATP to AMP + PPi
Glutamine synthetase is a central
control point in nitrogen
metabolism
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Gln is an amino donor for many biosynthetic products and
also a storage compound for excess ammonia.
Mammalian glutamine synthetase is activated by
ketoglutarate.
Bacterial glutamine synthetase has more complicated
regulation.
12 identical subunits, 469-aa, D6 symmetry.
Regulated by different effectors and covalent modification.
Figure 26-55a
X-Ray structure of S.
typhimurium glutamine synthetase. (a) View down the
6-fold axis showing only the six subunits of the upper
ring.
Active sites shown w/
Mn2+ ions (Mg2+)
Adenylation site is
indicated in yellow
(Tyr)
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ADP is shown in
cyan and
phosphinothricin is
shown (Glu inhibitor)
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Figure 26-55b
Side view of glutamine
synthetase along one of the enzyme’s 2-fold axes
showing only the eight nearest subunits.
Glutamine synthetase regulation
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9 feedback inhibitors control the activity of bacterial glutamine
synthetase
His, Trp, carbamoyl phosphate, glucosamine-6-phosphate,
AMP and CTP-pathways leading away from Gln
Ala, Ser, Gly-reflect cell’s N level
Ala, Ser, Gly, are competitive with Glu for the binding site.
AMP and CTP are competitive with the ATP binding site.
Glutamine synthetase regulation
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E. coli glutmine synthetase is covalently modified by adenylation of
a Tyr.
Increases susceptiblity to feedback inhibition and decreases activity
dependent on adenylation.
Adenylation and deadenylation are catalyzed by adenylyltransferase in
complex with a tetrameric regulatory protein, PII.
Adensyltransferase deadenylates glutamine synthetase when PII is
uridylated.
Adenylates glutamine synthetase when PII lacks UM residues.
PII uridylation depends on the activities of a uridylyltransferase and
uridylyl-removing enzyme that hydrolyzes uridylyl groups.
Glutamine synthetase regulation
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Uridylyltransferase is activated by -ketoglutarate
and ATP.
Uridylyltransferase is inhibited by glutamine and Pi.
Uridylyl-removing enzyme is insensitive to these
compounds.
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Figure 26-56 The
regulation of bacterial
glutamine synthetase.
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Figure 26-57 The
biosynthesis of the
“glutamate family” of
amino acids: arginine,
ornithine, and proline.
Conversion of Glu to Pro
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Involves reduction of the -carboxyl group to an aldehyde
followed for the formation of an internal Schiff base. This is
reduced to make Pro.
Proline synthesis
1. -glutamyl kinase
2. Dehydrogenase
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3. Nonenzymatic
4. Pyrroline-5carboxylate
reductase
Glutamate is the precursor for
Proline, Ornithine, and Arginine
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E. coli pathway from Gln to ornithine and Arg involves ATP-driven reduction of
the glutamate gamma carboxyl group to an aldehyde (N-acetylglutamate-5semialdehyde).
Spontaneous cyclization is prevented by acetylation of amino group by Nacetylglutamate synthase.
N-acetylglutamate-5-semialdehyde is converted to amine by transamination.
Hydrolysis of protecting group yields ornithine which can be converted to arginine.
In humans it is direct from glutamate-5-semialdehyde to ornithine by ornithine-aminotransferase
Arginine synthesis
5.glutamyl kinase
6. Acetylglutamate kinase
7. N-acetyl--glutamyl
phosphate
dehydrogense
8. N-acetylornithine-aminotransferase
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9. Acetylornithine
deacetylase
10. ornithine-aminotransferase
11. Urea cycle to arginine
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Figure 26-58 The conversion of
glycolytic intermediate 3phosphoglycerate to serine.
1. Conversion of 3phosphoglycerate’s 2-OH
group to a ketone
2. Transamination of 3phosphohydroxypyruvate
to 3-phosphoserine
3. Hydrolysis of
phosphoserine to make
Ser.
Serine is the precursor for Gly
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1.
2.
Ser can act in glycine synthesis in two ways:
Direct conversion of serine to glycine by hydroxymethyl transferase in
reverse (also yields N5, N10-methylene-THF)
Condensation of the N5, N10-methylene-THF with CO2 and NH4+ by the glycine
cleavage system
Cys derived from Ser
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In animals, Cys is derived from Ser and homocysteine
(breakdown product of Met).
The -SH group is derived from Met, so Cys can be
considered essential.
Methionine adenosyltransferase
2.
Methyltransferase
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Adenosylhomocysteinase
4.
Methionine synthase (B12)
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Cystathionine -synthase (PLP)
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Cystathionine -synthase (PLP)
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-ketoacid dehydrogenase
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Propionyl-CoA carboxylase (biotin)
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Methylmalonyl-CoA racemase
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Methylmalonyl-CoA mutase
11.
Glycine cleavage system or serine
hydroxymethyltransferase
12.
N5,N10-methylene-tetrahydrofolate
reductase (coenzyme B12 and
FAD)
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1.
Cys derived from Ser
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In plants and microorganisms, Cys is synthesized from Ser in two step reaction.
Reaction 1: activation of Ser -OH group by converting to O-acetylserine.
Reaction 2: displacement of the acetate by sulfide.
Sulfide is derived fro man 8-electron reduction reaction.
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Figure 26-59a
Cysteine
biosynthesis. (a) The
synthesis of cysteine from
serine in plants and
microorganisms.
Figure 26-59b
Cysteine
biosynthesis. (b) The 8electron reduction of
sulfate to sulfide in E. coli.
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1. Sulfate activation by
ATP sulfuylase and
adeosine-5’phosphosulfate (APS)
kinase
2. Sulfate reduced to sulfite
by 3’phosphoadenosine-5’phosphosulfate (PAPS)
reductase
3. Sulfite to sulfide by
sulfite reductase
Biosynthesis of essential amino
acids
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Pathways only present in microorganisms and plants.
Derived from metabolic precursors.
Usually involve more steps than nonessential amino acids.
Biosynthesis of Lys, Met, Thr
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First reaction is catalyzed by aspartokinase which converts aspartate to
apartyl--phosphate.
Each pathway is independently controlled.
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Figure 26-60 The
biosynthesis of the
“aspartate family” of
amino acids: lysine,
methionine, and
threonine.
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Figure 26-61 The
biosynthesis of the
“pyruvate family” of
amino acids:
isoleucine, leucine,
and valine.
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Figure 26-62 The
biosynthesis of
chorismate, the
aromatic amino acid
precursor.
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Figure 26-63
The
biosynthesis of
phenylalanine,
tryptophan, and
tyrosine from
chorismate.
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Figure 26-64 A ribbon diagram of the bifunctional
enzyme tryptophan synthase from S. typhimurium
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Figure 26-65 The
biosynthesis of
histidine.