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Literature summary for 1.14.20.1 extracted from
- Engineering Streptomyces clavuligerus deacetoxycephalosporin C synthase for optimal ring expansion activity toward penicillin G (2003), Appl. Environ. Microbiol., 69, 2306-2312.
Application
| Application | Comment | Organism |
|---|---|---|
| synthesis | enzyme produces the 7-aminodeacetoxycephalosporanic acid precursor utilized in industrial applications, optimization of enzyme activity for activity with the substrate penicillin G | Streptomyces clavuligerus |
Cloned(Commentary)
| Cloned (Comment) | Organism |
|---|---|
| expression of wild-type and mutant enzymes in Escherichia coli strain ESS | Streptomyces clavuligerus |
Protein Variants
| Protein Variants | Comment | Organism |
|---|---|---|
| C281Y | random mutagenesis, exchange of residue surrounding the substrate, 2-6fold increased activity compared to the wild-type enzyme | Streptomyces clavuligerus |
| G79E | random mutagenesis, exchange of residue surrounding the substrate, 2-6fold increased activity compared to the wild-type enzyme | Streptomyces clavuligerus |
| I305L | site-directed mutagenesis, exchange of residue surrounding the substrate, 6-14fold increased activity compared to the wild-type enzyme | Streptomyces clavuligerus |
| I305M | site-directed mutagenesis, exchange of residue surrounding the substrate, 6-14fold increased activity compared to the wild-type enzyme | Streptomyces clavuligerus |
| N304K | site-directed mutagenesis, exchange of residue surrounding the substrate, 6-14fold increased activity compared to the wild-type enzyme | Streptomyces clavuligerus |
| V275I | random mutagenesis, exchange of residue surrounding the substrate, 2-6fold increased activity compared to the wild-type enzyme | Streptomyces clavuligerus |
| V275I/C281Y/I305M | site-directed mutagenesis, exchange of residue surrounding the substrate, over 32fold increased activity compared to the wild-type enzyme | Streptomyces clavuligerus |
| V275I/I305M | site-directed mutagenesis, exchange of residue surrounding the substrate, 32fold increased activity compared to the wild-type enzyme | Streptomyces clavuligerus |
KM Value [mM]
| KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
|---|---|---|---|---|---|
| 0.004 | - |
Penicillin N | pH 6.5, 30°C, recombinant mutant N304K | Streptomyces clavuligerus |  |
| 0.006 | - |
Penicillin N | pH 6.5, 30°C, recombinant mutants C281Y and I305L | Streptomyces clavuligerus | |
| 0.009 | - |
Penicillin N | pH 6.5, 30°C, recombinant mutant G79E | Streptomyces clavuligerus | |
| 0.012 | - |
Penicillin N | pH 6.5, 30°C, recombinant mutants V275I and I305M | Streptomyces clavuligerus | |
| 0.013 | - |
Penicillin N | pH 6.5, 30°C, recombinant mutant V275I/I305M | Streptomyces clavuligerus | |
| 0.014 | - |
Penicillin N | pH 6.5, 30°C, recombinant wild-type enzyme | Streptomyces clavuligerus | |
| 0.22 | - |
penicillin G | pH 6.5, 30°C, recombinant mutant N304K | Streptomyces clavuligerus | |
| 0.25 | - |
penicillin G | pH 6.5, 30°C, recombinant mutant V275I/I305M | Streptomyces clavuligerus | |
| 0.66 | - |
penicillin G | pH 6.5, 30°C, recombinant mutant I305L | Streptomyces clavuligerus | |
| 0.68 | - |
penicillin G | pH 6.5, 30°C, recombinant mutant C281Y | Streptomyces clavuligerus | |
| 0.75 | - |
penicillin G | pH 6.5, 30°C, recombinant mutants G79E and I305M | Streptomyces clavuligerus | |
| 1.68 | - |
penicillin G | pH 6.5, 30°C, recombinant mutant V275I | Streptomyces clavuligerus | |
| 2.58 | - |
penicillin G | pH 6.5, 30°C, recombinant wild-type enzyme | Streptomyces clavuligerus | |
Natural Substrates/ Products (Substrates)
| Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
|---|---|---|---|---|---|---|
| penicillin G + 2-oxoglutarate + O2 | Streptomyces clavuligerus | - |
phenylacetyl-7-aminodeacetoxycephalosporanic acid + succinate + CO2 + H2O | product is the precursor of 7-aminodeacetoxycephalosporanic acid, which is used in industrial applications | ir | |
| penicillin N + 2-oxoglutarate + O2 | Streptomyces clavuligerus | - |
deacetoxycephalosporin C + succinate + CO2 + H2O | - |
ir | |
Organism
| Organism | UniProt | Comment | Textmining |
|---|---|---|---|
| Streptomyces clavuligerus | - |
gene cefE | - |
Purification (Commentary)
| Purification (Comment) | Organism |
|---|---|
| recombinant wild-type and mutant enzymes from Escherichia coli strain ESS by ion exchange chromatography and gel filtration | Streptomyces clavuligerus |
Specific Activity [micromol/min/mg]
| Specific Activity Minimum [µmol/min/mg] | Specific Activity Maximum [µmol/min/mg] | Comment | Organism |
|---|---|---|---|
| additional information | - |
activity of recombinant wild-type and mutant enzymes with substrate penicillin G and penicillin N | Streptomyces clavuligerus |
Substrates and Products (Substrate)
| Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
|---|---|---|---|---|---|---|
| penicillin G + 2-oxoglutarate + O2 | - |
Streptomyces clavuligerus | phenylacetyl-7-aminodeacetoxycephalosporanic acid + succinate + CO2 + H2O | - |
ir | |
| penicillin G + 2-oxoglutarate + O2 | - |
Streptomyces clavuligerus | phenylacetyl-7-aminodeacetoxycephalosporanic acid + succinate + CO2 + H2O | product is the precursor of 7-aminodeacetoxycephalosporanic acid, which is used in industrial applications | ir | |
| penicillin N + 2-oxoglutarate + O2 | - |
Streptomyces clavuligerus | deacetoxycephalosporin C + succinate + CO2 + H2O | - |
ir | |
Synonyms
| Synonyms | Comment | Organism |
|---|---|---|
| DAOCS | - |
Streptomyces clavuligerus |
| deacetoxycephalosporin C synthase | - |
Streptomyces clavuligerus |
Temperature Optimum [°C]
| Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
|---|---|---|---|
| 30 | - |
assay at | Streptomyces clavuligerus |
Turnover Number [1/s]
| Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
|---|---|---|---|---|---|
| 0.032 | - |
penicillin G | pH 6.5, 30°C, recombinant mutant G79E | Streptomyces clavuligerus | |
| 0.045 | - |
penicillin G | pH 6.5, 30°C, recombinant wild-type enzyme | Streptomyces clavuligerus | |
| 0.05 | - |
penicillin G | pH 6.5, 30°C, recombinant mutant V275I | Streptomyces clavuligerus | |
| 0.056 | - |
penicillin G | pH 6.5, 30°C, recombinant mutant N304K | Streptomyces clavuligerus | |
| 0.074 | - |
penicillin G | pH 6.5, 30°C, recombinant mutant C281Y | Streptomyces clavuligerus | |
| 0.076 | - |
penicillin G | pH 6.5, 30°C, recombinant mutant I305L | Streptomyces clavuligerus | |
| 0.145 | - |
penicillin G | pH 6.5, 30°C, recombinant mutant I305M | Streptomyces clavuligerus | |
| 0.146 | - |
penicillin G | pH 6.5, 30°C, recombinant mutant V275I/I305M | Streptomyces clavuligerus | |
| 0.178 | - |
Penicillin N | pH 6.5, 30°C, recombinant mutant G79E | Streptomyces clavuligerus | |
| 0.252 | - |
Penicillin N | pH 6.5, 30°C, recombinant mutant V275I | Streptomyces clavuligerus | |
| 0.273 | - |
Penicillin N | pH 6.5, 30°C, recombinant mutant C281Y | Streptomyces clavuligerus | |
| 0.284 | - |
Penicillin N | pH 6.5, 30°C, recombinant mutant I305L | Streptomyces clavuligerus | |
| 0.307 | - |
Penicillin N | pH 6.5, 30°C, recombinant wild-type enzyme | Streptomyces clavuligerus | |
| 0.31 | - |
Penicillin N | pH 6.5, 30°C, recombinant mutant I305M | Streptomyces clavuligerus | |
| 0.316 | - |
Penicillin N | pH 6.5, 30°C, recombinant mutant V275I/I305M | Streptomyces clavuligerus | |
| 0.366 | - |
Penicillin N | pH 6.5, 30°C, recombinant mutant N304K | Streptomyces clavuligerus | |
pH Optimum
| pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
|---|---|---|---|
| 6.5 | - |
assay at, substrate penicillin G | Streptomyces clavuligerus |
| 7.5 | - |
assay at, substrate penicillin N | Streptomyces clavuligerus |
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