Cloned (Comment) | Organism |
---|---|
recombinant expression of wild-type and mutant enzymes in Escherichia coli strain BL21(DE3) | Symbiobacterium thermophilum |
Protein Variants | Comment | Organism |
---|---|---|
H227C | site-directed saturation mutagenesis, the mutant shows 15.1fold increased activity with phenylpyruvate compared to the wild-type enzyme | Symbiobacterium thermophilum |
H227V | site-directed saturation mutagenesis, the mutant shows 35.1fold increased activity with phenylpyruvate compared to the wild-type enzyme | Symbiobacterium thermophilum |
additional information | in order to enlarge the substrate binding pocket of the meso-diaminopimelate dehydrogenase from Symbiobacterium thermophilum to accommodate larger 2-keto acids, e.g. phenylalanine, four amino acid residues, Phe146, Thr171, Arg181, and His227, are targeted for site saturation mutagenesis | Symbiobacterium thermophilum |
R181F | site-directed saturation mutagenesis, the mutant shows 6.4fold increased activity with phenylpyruvate compared to the wild-type enzyme | Symbiobacterium thermophilum |
R181F/H227V | site-directed saturation mutagenesis, the mutant shows 19.3fold increased activity with phenylpyruvate compared to the wild-type enzyme | Symbiobacterium thermophilum |
T171P | site-directed saturation mutagenesis, the mutant shows 2.2fold increased activity with phenylpyruvate compared to the wild-type enzyme | Symbiobacterium thermophilum |
T171S | site-directed saturation mutagenesis, the mutant shows 2.8fold increased activity with phenylpyruvate compared to the wild-type enzyme | Symbiobacterium thermophilum |
T171S/H227V | site-directed saturation mutagenesis, the mutant shows 10.6fold increased activity with phenylpyruvate compared to the wild-type enzyme | Symbiobacterium thermophilum |
T171S/R181F | site-directed saturation mutagenesis, the mutant shows 4.0fold increased activity with phenylpyruvate compared to the wild-type enzyme | Symbiobacterium thermophilum |
T171S/R181F/H227V | site-directed saturation mutagenesis, the mutant shows 14.6fold increased activity with phenylpyruvate compared to the wild-type enzyme | Symbiobacterium thermophilum |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
11 | - |
phenylpyruvate | pH 8.5, 30°C, recombinant mutant T171S/R181F/H227V | Symbiobacterium thermophilum | |
11.1 | - |
phenylpyruvate | pH 8.5, 30°C, recombinant mutant R181F | Symbiobacterium thermophilum | |
12.5 | - |
phenylpyruvate | pH 8.5, 30°C, recombinant wild-type enzyme | Symbiobacterium thermophilum | |
13.9 | - |
phenylpyruvate | pH 8.5, 30°C, recombinant mutant T171S/H227V | Symbiobacterium thermophilum | |
13.9 | - |
phenylpyruvate | pH 8.5, 30°C, recombinant mutant T171S/R181F | Symbiobacterium thermophilum | |
15.5 | - |
phenylpyruvate | pH 8.5, 30°C, recombinant mutant H227C | Symbiobacterium thermophilum | |
15.8 | - |
phenylpyruvate | pH 8.5, 30°C, recombinant mutantT171P | Symbiobacterium thermophilum | |
19.6 | - |
phenylpyruvate | pH 8.5, 30°C, recombinant mutant T171S | Symbiobacterium thermophilum | |
20.8 | - |
phenylpyruvate | pH 8.5, 30°C, recombinant mutant R181F/H227V | Symbiobacterium thermophilum | |
24.3 | - |
phenylpyruvate | pH 8.5, 30°C, recombinant mutant H227V | Symbiobacterium thermophilum |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
meso-2,6-diaminoheptanedioate + H2O + NADP+ | Symbiobacterium thermophilum | - |
L-2-amino-6-oxoheptanedioate + NH3 + NADPH + H+ | - |
r |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Symbiobacterium thermophilum | Q67PI3 | - |
- |
Purification (Comment) | Organism |
---|---|
recombinant wild-type and mutant enzymes from Escherichia coli strain Bl21(DE3) | Symbiobacterium thermophilum |
Specific Activity Minimum [µmol/min/mg] | Specific Activity Maximum [µmol/min/mg] | Comment | Organism |
---|---|---|---|
0.07 | - |
recombinant wild-type enzyme, pH 8.5, 30°C | Symbiobacterium thermophilum |
0.15 | - |
recombinant mutant T171P, pH 8.5, 30°C | Symbiobacterium thermophilum |
0.19 | - |
recombinant mutant T171S, pH 8.5, 30°C | Symbiobacterium thermophilum |
0.28 | - |
recombinant mutant T171S/R181F, pH 8.5, 30°C | Symbiobacterium thermophilum |
0.44 | - |
recombinant mutant R181F, pH 8.5, 30°C | Symbiobacterium thermophilum |
0.74 | - |
recombinant mutant T171S/H227V, pH 8.5, 30°C | Symbiobacterium thermophilum |
1.02 | - |
recombinant mutant T171S/R181F/H227V, pH 8.5, 30°C | Symbiobacterium thermophilum |
1.03 | - |
recombinant mutant H227C, pH 8.5, 30°C | Symbiobacterium thermophilum |
1.35 | - |
recombinant mutant R181F/H227V, pH 8.5, 30°C | Symbiobacterium thermophilum |
2.39 | - |
recombinant mutant H227V, pH 8.5, 30°C | Symbiobacterium thermophilum |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
meso-2,6-diaminoheptanedioate + H2O + NADP+ | - |
Symbiobacterium thermophilum | L-2-amino-6-oxoheptanedioate + NH3 + NADPH + H+ | - |
r | |
phenylpyruvate + NH3 + NADPH + H+ | - |
Symbiobacterium thermophilum | phenylalanine + NADP+ | - |
? |
Synonyms | Comment | Organism |
---|---|---|
meso-DAPDH | - |
Symbiobacterium thermophilum |
meso-diaminopimelate dehydrogenase | - |
Symbiobacterium thermophilum |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
30 | - |
assay at | Symbiobacterium thermophilum |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.11 | - |
phenylpyruvate | pH 8.5, 30°C, recombinant wild-type enzyme | Symbiobacterium thermophilum | |
0.53 | - |
phenylpyruvate | pH 8.5, 30°C, recombinant mutantT171P | Symbiobacterium thermophilum | |
0.7 | - |
phenylpyruvate | pH 8.5, 30°C, recombinant mutant T171S/R181F | Symbiobacterium thermophilum | |
0.88 | - |
phenylpyruvate | pH 8.5, 30°C, recombinant mutant T171S | Symbiobacterium thermophilum | |
1.07 | - |
phenylpyruvate | pH 8.5, 30°C, recombinant mutant R181F | Symbiobacterium thermophilum | |
2.09 | - |
phenylpyruvate | pH 8.5, 30°C, recombinant mutant T171S/R181F/H227V | Symbiobacterium thermophilum | |
2.37 | - |
phenylpyruvate | pH 8.5, 30°C, recombinant mutant T171S/H227V | Symbiobacterium thermophilum | |
2.48 | - |
phenylpyruvate | pH 8.5, 30°C, recombinant mutant H227C | Symbiobacterium thermophilum | |
3.43 | - |
phenylpyruvate | pH 8.5, 30°C, recombinant mutant R181F/H227V | Symbiobacterium thermophilum | |
3.98 | - |
phenylpyruvate | pH 8.5, 30°C, recombinant mutant H227V | Symbiobacterium thermophilum |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
8.5 | - |
assay at | Symbiobacterium thermophilum |
Cofactor | Comment | Organism | Structure |
---|---|---|---|
NADP+ | - |
Symbiobacterium thermophilum | |
NADPH | - |
Symbiobacterium thermophilum |
General Information | Comment | Organism |
---|---|---|
additional information | molecular dynamic simulation analysis of wild-type and variant H227V enzyme with the imine intermediate as the substrate, overview | Symbiobacterium thermophilum |