Cloned (Comment) | Organism |
---|---|
open reading frames for P450cam mutants including a 6 x His carboxyl-terminal tag cloned into the pCW(Ori+) expression vector using the NdeI and XbaI restriction sites, expressed in Escherichia coli | Pseudomonas putida |
Protein Variants | Comment | Organism |
---|---|---|
T252A | non-productive H2O2 generation is dominant, does not oxidize camphor, substrate binding affinity is similar to that of the wild-type enzyme | Pseudomonas putida |
T252N | has comparable turnover number but higher Km value relative to the wild-type enzyme, due to a decrease in the camphor binding affinity, non-productive H2O2 generation is negligible | Pseudomonas putida |
T252N/V253T | has comparable turnover number but higher Km value relative to the wild-type enzyme, due to a decrease in the camphor binding affinity, non-productive H2O2 generation is negligible | Pseudomonas putida |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.03 | - |
(+)-Camphor | wild-type | Pseudomonas putida | |
0.096 | - |
(+)-Camphor | mutant T252N | Pseudomonas putida | |
0.388 | - |
(+)-Camphor | mutant T252N/V253T | Pseudomonas putida |
Metals/Ions | Comment | Organism | Structure |
---|---|---|---|
Fe2+ | Asn252 can stabilize the ferric hydroperoxy intermediate, preventing premature release of H2O2 and enabling addition of the second proton to the distal oxygen to generate the catalytic ferryl species | Pseudomonas putida |
Molecular Weight [Da] | Molecular Weight Maximum [Da] | Comment | Organism |
---|---|---|---|
47000 | - |
gel filtration | Pseudomonas putida |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Pseudomonas putida | P00183 | - |
- |
Purification (Comment) | Organism |
---|---|
soluble proteins separated by ultracentrifugation and purified using Ni2+-nitrilotriacetate column and gel filtration | Pseudomonas putida |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
(+)-camphor + O2 + reduced putidaredoxin | - |
Pseudomonas putida | (+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O | - |
? |
Synonyms | Comment | Organism |
---|---|---|
CYP101 | - |
Pseudomonas putida |
cytochrome p450cam | - |
Pseudomonas putida |
P450cam | - |
Pseudomonas putida |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.9 | - |
(+)-Camphor | mutant T252N | Pseudomonas putida | |
2.067 | - |
(+)-Camphor | wild-type | Pseudomonas putida | |
2.75 | - |
(+)-Camphor | mutant T252N/V253T | Pseudomonas putida |
Cofactor | Comment | Organism | Structure |
---|---|---|---|
putidaredoxin | - |
Pseudomonas putida |