Application | Comment | Organism |
---|---|---|
synthesis | the enzyme KcPGA is quite resilient to harsh conditions and is easier to immobilize for the industrial hydrolysis of natural penicillins to generate the 6-aminopenicillin nucleus, which is the starting material for semisynthetic antibiotic production | Kluyvera cryocrescens |
Cloned (Comment) | Organism |
---|---|
gene pac, recombinant expression of N-terminally His-tagged wild-type and mutant enzymes in Escherichia coli strain BL21 (DE3) pLysS | Kluyvera cryocrescens |
Crystallization (Comment) | Organism |
---|---|
purified recombinant enzyme, sitting drop vapour diffusion method, mixing of 300 nl of 45 mg/ml protein solution with 0.1 ml of precipitant solution containing 30% w/v PEG 4000, 50 mM sodium cacodylate pH 5.6, 0.5 M potassium thiocyanate, 3-4 days, mixing of 500 nl of protein and of precipitant solutions each results in crystals that grow in a week under a wide range of pH conditions using 50 mM sodium cacodylate buffer, method optimization, X-ray diffraction structure determination and analysis at 2.5-3.5 A resolution, molecular replacement method | Kluyvera cryocrescens |
Protein Variants | Comment | Organism |
---|---|---|
S290G | site-directed mutagenesis of the internal proteolytic cleavage site of the pro-enzyme | Kluyvera cryocrescens |
Localization | Comment | Organism | GeneOntology No. | Textmining |
---|
Molecular Weight [Da] | Molecular Weight Maximum [Da] | Comment | Organism |
---|---|---|---|
92000 | - |
1 * 92000, pro-enzyme, SDS-PAGE | Kluyvera cryocrescens |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
penicillin G + H2O | Kluyvera cryocrescens | - |
phenylacetate + 6-aminopenicillanate | - |
? | |
penicillin G + H2O | Kluyvera cryocrescens DMSZ 2660 | - |
phenylacetate + 6-aminopenicillanate | - |
? |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Kluyvera cryocrescens | P07941 | gene pac | - |
Kluyvera cryocrescens DMSZ 2660 | P07941 | gene pac | - |
Posttranslational Modification | Comment | Organism |
---|---|---|
proteolytic modification | the enzyme is synthesized as a single-chain inactive pro-enzyme, which upon autocatalytic processing becomes an active heterodimer of alpha and beta chains. Cleavage of the Thr289-Ser290 bond leads to the unveiling of the primary amine group of Serbeta1 (Ser290 of the precursor), creating the active centre in mature enzyme | Kluyvera cryocrescens |
Purification (Comment) | Organism |
---|---|
recombinant N-terminally His-tagged wild-type and mutant enzymes to homogeneity from Escherichia coli strain BL21 (DE3) pLysS by nickel affinity chromatography and gel filtration | Kluyvera cryocrescens |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
penicillin G + H2O | - |
Kluyvera cryocrescens | phenylacetate + 6-aminopenicillanate | - |
? | |
penicillin G + H2O | - |
Kluyvera cryocrescens DMSZ 2660 | phenylacetate + 6-aminopenicillanate | - |
? |
Subunits | Comment | Organism |
---|---|---|
heterodimer | - |
Kluyvera cryocrescens |
monomer | 1 * 92000, pro-enzyme, SDS-PAGE | Kluyvera cryocrescens |
Synonyms | Comment | Organism |
---|---|---|
penicillin G acylase | - |
Kluyvera cryocrescens |
PGA | - |
Kluyvera cryocrescens |