Penicillin acyltransferase in Penicillium chrysogenum.
Isotopic exchange of (35)S between penicillins and 6-amino-penicillanic acid (6-APA) was observed in cell-free extracts of Penicillium chrysogenum. Sulfhydryl-containing compounds were required for activity. Dithiothreitol, dithioerythritol, mercaptoethanol, and glutathione served as activators. The acyltransferase was purified threefold by adsorption on calcium phosphate gel at pH 6 and elution at pH 8. The partially purified enzyme showed maximal activity at pH 8. The enzyme was stable at 25 C for at least 30 min at pH 8. Dissociable inhibitors or activators, other than the sulfhydryl-containing compounds, could not be demonstrated in the enzyme preparation. An apparent Michaelis constant of 1.5 +/- 0.5 mm was determined for penicillin G at a 6-APA concentration of 5 mm. The enzyme did not appear to possess penicillin amidase activity. The exchange mechanism probably involves an acyl-enzyme intermediate. Penicillins V, G, K, X, and dihydro F showed isotopic exchange with (35)S-6-APA. Penicillin N, methylpenicillin, and phenyl-penicillin did not show exchange. The level of acyltransferase in P. chrysogenum 51-20F3 was measured at times during the fermentation. The level of activity increased threefold between 40 and 55 hr, remaining high until about 90 hr.
D000217:Acyltransferases / Q000378:metabolism*; D000581:Amidohydrolases / Q000378:metabolism; D000596:Amino Acids / Q000378:metabolism; D001230:Aspergillus / Q000201:enzymology; D002474:Cell-Free System; D005978:Glutathione / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008623:Mercaptoethanol / Q000494:pharmacology; D010400:Penicillin G / Q000378:metabolism; D010404:Penicillin V / Q000378:metabolism; D010406:Penicillins / Q000378:metabolism*; D010407:Penicillium / Q000201:enzymology*; D010648:Phenylacetates / Q000494:pharmacology; D013438:Sulfhydryl Compounds / Q000494:pharmacology; D013460:Sulfur Isotopes
D000596:Amino Acids; D010406:Penicillins; D010648:Phenylacetates; D013438:Sulfhydryl Compounds; D013460:Sulfur Isotopes; D008623:Mercaptoethanol; D000217:Acyltransferases; D000581:Amidohydrolases; D005978:Glutathione; D010400:Penicillin G; D010404:Penicillin V