Cloned (Comment) | Organism |
---|---|
mutant enzyme S96A is produced in cytoplasm of Escherichia coli. Mutant enzymes K38H, C98A and C98N are produced in culture medium of Streptomyces lividans | Streptomyces sp. |
Crystallization (Comment) | Organism |
---|---|
hanging drop vapor diffusion method. Mutant enzymes S96A, K38H, C98A and C98N are produced in culture medium of Streptomyces lividans | Streptomyces sp. |
Protein Variants | Comment | Organism |
---|---|---|
C98A | half-life at 72°C increases to 13 min, compared to 10 min for the wild-type enzyme. Half-life at 60°C increases to 220 min compared to 170 min for the wild-type enzyme. The value of the second-order acylation rate constant is not very different from those of the wild-type enzyme with the exception of the cephalotin, oxacillin and piperacillin values which are increased 5-10fold | Streptomyces sp. |
C98N | half-life at 72°C decreases to 6 min, compared to 10 min for the wild-type enzyme. Half-life at 60°C decreases to 30 min compared to 170 min for the wild-type enzyme. The acylation rate constants by most of the beta-lactams are increased 10-70fold compared to the wild-type enzyme. The deacylation rate constant values als increases | Streptomyces sp. |
K38H | half-life at 72°C decreases to 3.5 min, compared to 10 min for the wild-type enzyme. The mutant enzyme fails to bind beta-lactams | Streptomyces sp. |
S96A | half-life at 72°C decreases to 8 min, compared to 10 min for the wild-type enzyme. Half-life at 60°C decreases to 120 min compared to 170 min for the wild-type enzyme. The mutant enzyme is inactive in terms of beta-lactam binding with the exception of cefoxitin, for which a residual activity is detected.The value of the second-order acylation rate constant is decreased by a factor of 3000, and that of the deacylation rate constant is decreased 120fold | Streptomyces sp. |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.22 | - |
benzoyl-Gly-thiolactate | pH 7.2, 37°C, wild-type enzyme | Streptomyces sp. | |
0.3 | - |
benzoyl-Gly-thiolactate | pH 7.2, 37°C, mutant enzyme C98A | Streptomyces sp. | |
0.35 | - |
Nalpha,Nepsilon-Diacetyl-L-Lys-D-Ala-D-Ala | above, pH 7.2, 37°C, mutant enzyme C98A | Streptomyces sp. | |
0.4 | - |
N-alpha-acetyl-L-Lys-D-Ala-thiolactate | pH 7.2, 37°C, wild-type enzyme | Streptomyces sp. | |
0.6 | - |
Nalpha-acetyl-L-Lys-D-Ala-thiolactate | pH 7.2, 37°C, mutant enzyme C98A | Streptomyces sp. | |
0.8 | - |
benzoyl-Gly-thioglycolate | pH 7.2, 37°C, mutant enzyme C98A | Streptomyces sp. | |
1 | - |
benzoyl-Gly-thiolactate | pH 7.2, 37°C, mutant enzyme K38H | Streptomyces sp. | |
1.2 | - |
phenylacetyl-D-Ala-thiolactate | pH 7.2, 37°C, mutant enzyme C98A | Streptomyces sp. | |
1.2 | - |
phenylacetyl-D-Ala-thiolactate | pH 7.2, 37°C, wild-type enzyme | Streptomyces sp. | |
1.3 | - |
benzoyl-D-Ala-thioglycolate | pH 7.2, 37°C, mutant enzyme C98A | Streptomyces sp. | |
1.5 | - |
benzoyl-D-Ala-thioglycolate | pH 7.2, 37°C, wild-type enzyme | Streptomyces sp. | |
2 | - |
Nalpha,Nepsilon-diacetyl-L-Lys-D-Ala-thiolactate | pH 7.2, 37°C, mutant enzyme C98A | Streptomyces sp. | |
2.4 | - |
phenylacetyl-D-Ala-thioglycolate | pH 7.2, 37°C, mutant enzyme K38H | Streptomyces sp. | |
3 | - |
phenylacetyl-D-Ala-thioglycolate | above, pH 7.2, 37°C, mutant enzyme C98A | Streptomyces sp. | |
3 | - |
phenylacetyl-D-Ala-thioglycolate | above, pH 7.2, 37°C, wild-type enzyme | Streptomyces sp. | |
3 | - |
Nalpha,Nepsilon-diacetyl-L-Lys-D-Ala-thiolactate | above, pH 7.2, 37°C, wild-type enzyme | Streptomyces sp. | |
3 | - |
benzoyl-Gly-thioglycolate | above, pH 7.2, 37°C, wild-type enzyme | Streptomyces sp. | |
6.2 | - |
Nalpha,Nepsilon-Diacetyl-L-Lys-D-Ala-D-Ala | pH 7.2, 37°C, wild-type enzyme | Streptomyces sp. | |
9.4 | - |
Nalpha-Acetyl-L-Lys-D-Ala-D-Ala | pH 7.2, 37°C, wild-type enzyme | Streptomyces sp. | |
30 | - |
Nalpha-Acetyl-L-Lys-D-Ala-D-Ala | pH 7.2, 37°C, mutant enzyme C98A | Streptomyces sp. |
Localization | Comment | Organism | GeneOntology No. | Textmining |
---|---|---|---|---|
cytoplasm | mutant enzyme S96A is produced in cytoplasm of Escherichia coli | Streptomyces sp. | 5737 | - |
extracellular | mutant enzymes K38H, C98A and C98N are produced in culture medium of Streptomyces lividans | Streptomyces sp. | - |
- |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Streptomyces sp. | P39042 | K15 | - |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
benzoyl-D-Ala-thioglycolate + H2O | - |
Streptomyces sp. | ? | - |
? | |
benzoyl-Gly-thioglycolate + H2O | - |
Streptomyces sp. | ? | - |
? | |
benzoyl-Gly-thiolactate + H2O | - |
Streptomyces sp. | ? | - |
? | |
N-alpha-acetyl-L-Lys-D-Ala-thiolactate + H2O | - |
Streptomyces sp. | ? | - |
? | |
Nalpha,Nepsilon-diacetyl-L-Lys-D-Ala-D-Ala + H2O | - |
Streptomyces sp. | Nalpha,Nepsilon-diacetyl-L-Lys-D-Ala + D-Ala | - |
? | |
Nalpha,Nepsilon-diacetyl-L-Lys-D-Ala-thiolactate + H2O | - |
Streptomyces sp. | ? | - |
? | |
Nalpha-Acetyl-L-Lys-D-Ala-D-Ala + H2O | - |
Streptomyces sp. | Nalpha-Acetyl-L-Lys-D-Ala + D-Ala | - |
? | |
phenylacetyl-D-Ala-thioglycolate + H2O | - |
Streptomyces sp. | ? | - |
? | |
phenylacetyl-D-Ala-thiolactate + H2O | - |
Streptomyces sp. | ? | - |
? |
Synonyms | Comment | Organism |
---|---|---|
DD-transpeptidase/penicillin-binding protein | - |
Streptomyces sp. |
Temperature Stability Minimum [°C] | Temperature Stability Maximum [°C] | Comment | Organism |
---|---|---|---|
60 | - |
half-lives: 120 min for mutant S96A, 220 min for mutant C98A, 30 min for mutant C98N and 170 min for the wild-type enzyme | Streptomyces sp. |
72 | - |
half-lives: 3.5 min for mutant K38H, 8 min for mutant S96A, 6 min for mutant C98A, 13 min for mutant enzyme C98A and 10 min for the wild-type enzyme | Streptomyces sp. |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.006 | - |
benzoyl-Gly-thiolactate | pH 7.2, 37°C, mutant enzyme K38H | Streptomyces sp. | |
0.015 | - |
phenylacetyl-D-Ala-thioglycolate | pH 7.2, 37°C, mutant enzyme K38H | Streptomyces sp. | |
0.017 | - |
benzoyl-Gly-thiolactate | pH 7.2, 37°C, wild-type enzyme | Streptomyces sp. | |
0.024 | - |
benzoyl-Gly-thioglycolate | above, pH 7.2, 37°C, wild-type enzyme | Streptomyces sp. | |
0.08 | - |
benzoyl-Gly-thiolactate | pH 7.2, 37°C, mutant enzyme C98A | Streptomyces sp. | |
0.08 | - |
benzoyl-Gly-thioglycolate | pH 7.2, 37°C, mutant enzyme C98A | Streptomyces sp. | |
0.1 | - |
N-alpha-acetyl-L-Lys-D-Ala-thiolactate | pH 7.2, 37°C, wild-type enzyme | Streptomyces sp. | |
0.11 | - |
benzoyl-D-Ala-thioglycolate | pH 7.2, 37°C, wild-type enzyme | Streptomyces sp. | |
0.13 | - |
Nalpha-Acetyl-L-Lys-D-Ala-D-Ala | pH 7.2, 37°C, mutant enzyme C98A | Streptomyces sp. | |
0.2 | - |
Nalpha,Nepsilon-Diacetyl-L-Lys-D-Ala-D-Ala | above, pH 7.2, 37°C, mutant enzyme C98A | Streptomyces sp. | |
0.3 | - |
benzoyl-D-Ala-thioglycolate | pH 7.2, 37°C, mutant enzyme C98A | Streptomyces sp. | |
0.3 | - |
Nalpha,Nepsilon-Diacetyl-L-Lys-D-Ala-D-Ala | pH 7.2, 37°C, wild-type enzyme | Streptomyces sp. | |
0.43 | - |
Nalpha,Nepsilon-diacetyl-L-Lys-D-Ala-thiolactate | above, pH 7.2, 37°C, wild-type enzyme | Streptomyces sp. | |
0.45 | - |
Nalpha-Acetyl-L-Lys-D-Ala-D-Ala | pH 7.2, 37°C, wild-type enzyme | Streptomyces sp. | |
0.5 | - |
phenylacetyl-D-Ala-thioglycolate | above, pH 7.2, 37°C, wild-type enzyme | Streptomyces sp. | |
0.6 | - |
phenylacetyl-D-Ala-thiolactate | pH 7.2, 37°C, wild-type enzyme | Streptomyces sp. | |
0.8 | - |
N-alpha-acetyl-L-Lys-D-Ala-thiolactate | pH 7.2, 37°C, mutant enzyme C98A | Streptomyces sp. | |
0.87 | - |
phenylacetyl-D-Ala-thioglycolate | above, pH 7.2, 37°C, mutant enzyme C98A | Streptomyces sp. | |
1.1 | - |
Nalpha,Nepsilon-diacetyl-L-Lys-D-Ala-thiolactate | pH 7.2, 37°C, mutant enzyme C98A | Streptomyces sp. | |
2.2 | - |
phenylacetyl-D-Ala-thiolactate | pH 7.2, 37°C, mutant enzyme C98A | Streptomyces sp. | |
6.08 | - |
phenylacetyl-D-Ala-thioglycolate | above, pH 7.2, 37°C, mutant enzyme C98A | Streptomyces sp. |