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2.6.1.1: aspartate transaminase

This is an abbreviated version!
For detailed information about aspartate transaminase, go to the full flat file.

Word Map on EC 2.6.1.1

Reaction

L-aspartate
+
2-oxoglutarate
=
oxaloacetate
 +
L-glutamate

Synonyms

2-oxoglutarate-glutamate aminotransferase, AAT, AAT-2, AAT-3, AAT3, aatA, AATase, aatB3, all2340, alr1039, alr2765, alr4853, aminotransferase, aspartate, aoa/coa, AsAT, Asp AT, aspartate alpha-ketoglutarate transaminase, aspartate aminotransferase, aspartate aminotransferase 1, aspartate aminotransferase A, aspartate AT, aspartate transaminase, aspartate, 2-oxoglutarate aminotransferase, aspartate-2-oxoglutarate transaminase, aspartate/(R)-cysteate:2-oxoglutarate aminotransferase, aspartate/tyrosine/phenylalanine pyridoxal-5'-phosphate-dependent aminotransferase, aspartate:2-oxoglutarate amino-transferase, aspartate:2-oxoglutarate aminotransferase, aspartic acid aminotransferase, aspartic aminotransferase, aspartyl aminotransferase, AspAT, AspATSs, aspB, aspC, AspT, AST, AST-Bb, AtPAT, bifunctional aspartate aminotransferase and glutamate/aspartate-prephenate aminotransferase, C-S lyase, CAA1, cAST, Cgl0240, class Ibeta AAT, EcAspAT, GL50803_91056, glutamate oxaloacetate transaminase, glutamate oxaloacetate transaminase 1, glutamate-oxalacetate aminotransferase, glutamate-oxalate transaminase, glutamate-oxaloacetate transaminase 1, glutamic oxalic transaminase, glutamic oxaloacetic transaminase, glutamic-aspartic aminotransferase, glutamic-aspartic transaminase, glutamic-oxalacetic transaminase, glutamic-oxaloacetic transaminase, GOT, GOT (enzyme), GOT1, GOT1L1, GOT2, KAT IV, L-aspartate aminotransferase, L-aspartate transaminase, L-aspartate-2-ketoglutarate aminotransferase, L-aspartate-2-oxoglutarate aminotransferase, L-aspartate-2-oxoglutarate-transaminase, L-aspartate-alpha-ketoglutarate transaminase, L-aspartate:2-oxoglutarate aminotransferase, L-aspartateartate aminotransferase, L-aspartic aminotransferase, L-AspAT, mitAAT, More, oxaloacetate transferase, oxaloacetate-aspartate aminotransferase, Pat, PfAspAT, plastid aspartate aminotransferase, prephenate aminotransferase, protein TT0402, PT-AAT, Rv3722c, Sar2028, SsAspAT, Tb11.02.2740, transaminase A

ECTree

     2 Transferases
         2.6 Transferring nitrogenous groups
             2.6.1 Transaminases
                2.6.1.1 aspartate transaminase

Engineering

Engineering on EC 2.6.1.1 - aspartate transaminase

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PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
A168G
site-directed mutagenesis, altered substrate binding kinetics compared to wild-type
E108K
site-directed mutagenesis, altered substrate binding kinetics compared to wild-type
K169S
site-directed mutagenesis, inactive with aspartate and glutamate
K169V
site-directed mutagenesis, inactive with glutamate, altered substrate binding kinetics compared to wild-type
K306A
site-directed mutagenesis, structure comparison with wild-type, overview. The alanine substitution of Lys306 prevents Schiff base formation with the cofactor, inactive mutant
R445G
site-directed mutagenesis, inactive mutant
T84V
site-directed mutagenesis, altered substrate binding kinetics compared to wild-type
T84V/K169V
site-directed mutagenesis, altered substrate binding kinetics compared to wild-type
D232N
mutant shows no activity
K270H
mutant shows no activity
R403Y
mutant shows no activity
D220A
mutation in pyridoxal phosphate-binding residue, almost complete loss of activity
K259A
mutation in pyridoxal phosphate-binding residue, almost complete loss of activity
K41A
mutation in substrate-binding residue, about 60% of wild-type activity
N191A
mutation in pyridoxal phosphate-binding residue, almost complete loss of activity
R144A
mutation in substrate-binding residue, about 25% of wild-type activity
R394A
mutation in substrate-binding residue, almost complete loss of activity
R39A
mutation in substrate-binding residue, almost complete loss of activity
S103A
mutation in pyridoxal phosphate-binding residue, about 70% of wild-type activity
S104A
mutation in pyridoxal phosphate-binding residue, about 70% of wild-type activity
S258A
mutation in pyridoxal phosphate-binding residue, about 80% of wild-type activity
Y142A
mutation in pyridoxal phosphate-binding residue, almost complete loss of activity
Y223A
mutation in pyridoxal phosphate-binding residue, almost complete loss of activity
Y73A
almost complete loss of activity
K259A
-
mutation in pyridoxal phosphate-binding residue, almost complete loss of activity
-
K41A
-
mutation in substrate-binding residue, about 60% of wild-type activity
-
S103A
-
mutation in pyridoxal phosphate-binding residue, about 70% of wild-type activity
-
S104A
-
mutation in pyridoxal phosphate-binding residue, about 70% of wild-type activity
-
Y73A
-
almost complete loss of activity
-
A12T/P13T/N34D/T109S/G261A/S285G/A293D/N297S
mutant enzyme has nearly the same ratio of kcat/Km(Phe) to kcat/Km(Asp) as that of wild-type enzyme. The additional mutation A293D compared to mutant enzyme A12T/P13T/N34D/T109S/G261A/S285G/N297S holds the Arg292 side chain away from the active site to allow increased specificity for phenylalanine over aspartate
E265K
-
site-directed mutagenesis, reduced kcat, reduced Km for L-aspartate and 2-oxoglutarate
E265Q
-
site-directed mutagenesis, reduced kcat, reduced Km for L-aspartate and 2-oxoglutarate
I33Q/Y214Q/R280Y
substitution of three active-site residues in Escherichia coli L-aspartateartate aminotransferase highly reduces kcat for transamination reaction. Ratio of L-cysteine sulfinate desulfinase to transaminase activity is increased by 100000fold in mutant. kcat for desulfination of L-cysteine sulfinate increases to 0.5/sec (from 0.05/sec in wild-type enzyme). kcat for beta-decarboxylation of L-aspartateartate increases from below 0.0001/sec to 0.07/sec
K258A
-
exchange of active site lysine for alanine, binds still pyridoxal 5'-phosphate or pyridoxamine 5'-phosphate, but not covalently, the latter forms when bound to the enzyme a covalent stable ketimine with 2-oxoglutarate and effects a beta-decarboxylation of oxaloacetate, followed by transamination to give the pyridoxal 5'-phosphate aldimine of alanine as a final product
K68E
-
site-directed mutagenesis, reduced kcat, reduced Km for L-aspartate and increased Km for 2-oxoglutarate
K68E/E265K
-
site-directed mutagenesis, charge exchange in the conserved intrasubunit salt bridge, reduced kcat, reduced Km for L-aspartate and 2-oxoglutarate
K68M
-
site-directed mutagenesis, reduced kcat, reduced Km for L-aspartate and increased Km for 2-oxoglutarate
K68M/E265Q
-
site-directed mutagenesis, introduction of a neutral amino acid pair in the conserved intrasubunit salt bridge, reduced kcat, reduced Km for L-aspartate and 2-oxoglutarate
N165Y/R316H/A381T/P399L
-
the mutant shows increased activity compared to the wild type enzyme
P138A
oligonucleotide-directed mutagenesis, no significant effect, unaltered compared to the wild-type
P138A/P195A
oligonucleotide-directed mutagenesis
P195A
oligonucleotide-directed mutagenesis, no significant effect, unaltered compared to the wild-type
R292E/L18H
-
12.9fold increase in specific activity towards L-Lys and 2-oxo-4-phenylbutanoic acid
T70F
-
nearly no activity with L-erythro-3-hydroxyaspartate, in opposite to the wild-type, altered reaction kinetics due to inability to stabilize the reaction quinonoid intermediate
V39F
mutant enzyme shows a more open conformation, and the aldimine pKa is lowered by 0.7 unit compared with the wild-type enzyme. The maleate-bound V39F enzyme shows the aldimine pKa 0.9 unit lower than that of the maleate-bound wild-type enzyme. The maleate-bound V39F enzyme shows an altered side-chain packing pattern in the 37–39 region, and the lack of repulsion between Gly38 carbonyl O and Tyr225 seems to be the cause of the reduced pKa value
V39F/N194A
mutation does not decrease the aldimine pKa, showing that the domain rotation controls the aldimine pKa via the Arg386-Asn194 pyridoxal 5'-phosphate linkage system
V39L
-
crystal structure analysis
V39L/K41Y/T47I/N69L/T109S/A293D/N297S
the additional mutation A293D compared to mutant enzyme V39L/K41Y/T47I/N69L/T109S/N297S holds the Arg292 side chain away from the active site to allow increased specificity for phenylalanine over aspartate
Y225R/R292K/R386A
the mutation converts its activity into an aspartate 4-decarboxylase
C166A
-
site-directed mutagenesis of isozyme mAspAT, decreased ability to undergo transition from the open to the closed conformation essential for the reaction mechanism, reduced reactivity with DTNB
C166S
-
site-directed mutagenesis of isozyme mAspAT, decreased ability to undergo transition from the open to the closed conformation essential for the reaction mechanism, reduced reactivity with DTNB
S256A
the mutant shows 3.51% activity compared to the wild type enzyme
T110A
the mutant shows 2.45% activity compared to the wild type enzyme
T110A/S256A
inactive
DELTA1-13
truncation of these noncatalytic residues reduces enzyme activity and a peptide containing these amino acids inhibits PfAspAT in vitro and in the lysate of cultured parasites
DELTA1-7
truncation of the first seven amino acids only minorly reduces enzymatic activity
K109S
site-directed mutagenesis, loss of activity towards acidic substrates, increased activity towards the neutral substrate alanine, increase in pKa value
K109V
site-directed mutagenesis, loss of activity towards acidic substrates, increased activity towards the neutral substrate alanine
K109S
Thermus thermophilus HB8 / ATCC 27634 / DSM 579
-
site-directed mutagenesis, loss of activity towards acidic substrates, increased activity towards the neutral substrate alanine, increase in pKa value
-
K109V
Thermus thermophilus HB8 / ATCC 27634 / DSM 579
-
site-directed mutagenesis, loss of activity towards acidic substrates, increased activity towards the neutral substrate alanine
-
K237A
additional information
-
construction of deletion mutant DELTA3-11mAspAT of isozyme mAspAT, enhanced thermostability, reduced kcat and Km, enhanced reactivity of Cys166 with DTNB