Purification and characterization of extracellular penicillin V acylase from Fusarium sp. SKF 235.
Penicillin V acylase from Fusarium sp. SKF 235 culture filtrate was purified to homogeneity. The enzyme was a glycoprotein and composed of single polypeptide chain with molecular weight of 83,200 Daltons. The pH and temperature optima were 6.5 and 55 degrees C, respectively. The KM for penicillin V was 10 mM but the enzyme was inhibited by penicillin V at concentrations above 50 mM. Products of reaction, 6-aminopenicillanic acid and phenoxyacetic acid inhibited the enzyme competitively and noncompetitively with Ki values of 18 mM and 45 mM, respectively. The enzyme specifically hydrolyzed penicillin V, cephalosporanic acid V and penicillin V sulphoxide. Other phenoxy acetyl amides studied were not hydrolysed. It is proposed that phenoxyacetyl moiety alone is not recognized by the penicillin V acylase and in addition, the beta-lactam structure contributes in formation of enzyme-substrate complex.
Sudhakaran
V K
VK
Research and Development, Hindustan Antibiotics Ltd., Pimpri, Pune, India.
Shewale
J G
JG
D000581:Amidohydrolases / Q000032:analysis / Q000378:metabolism; D001667:Binding, Competitive; D002511:Cephalosporins / Q000378:metabolism; D003470:Culture Media; D004591:Electrophoresis, Polyacrylamide Gel; D004791:Enzyme Inhibitors / Q000138:chemical synthesis / Q000378:metabolism*; D005670:Fusarium / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D008970:Molecular Weight; D010397:Penicillanic Acid / Q000031:analogs & derivatives / Q000737:chemistry; D010399:Penicillin Amidase / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000302:isolation & purification*; D010404:Penicillin V / Q000031:analogs & derivatives / Q000378:metabolism; D010642:Phenoxyacetates / Q000737:chemistry; D013379:Substrate Specificity; D013696:Temperature
D002511:Cephalosporins; D003470:Culture Media; D004791:Enzyme Inhibitors; D010642:Phenoxyacetates; C011325:penicillin V sulfoxide; D010397:Penicillanic Acid; D000581:Amidohydrolases; D010399:Penicillin Amidase; C019665:amidase; C004408:aminopenicillanic acid; D010404:Penicillin V