title: Partial purification and characterization of isopenicillin N epimerase activity from Streptomyces clavuligerus.
abstract: Epimerase activity, which converts isopenicillin N to penicillin N, has been partially purified from cell-free extracts of Streptomyces clavuligerus. No stimulating cofactors of this activity were found, and neither EDTA nor anaerobic incubation caused significant inhibition of activity. Although pyridoxal phosphate did not stimulate epimerase activity, the presence of this cofactor was necessary for the stabilization of enzymic activity during the purification process. Epimerase activity was purified 35.5-fold by a combination of salt precipitation, gel filtration, and ion exchange chromatography. Gel filtration indicated that the epimerase has a molecular weight of 60 000 and sodium dodecyl sulphate-polyacrylamide gel electrophoresis of the 35.5-fold purified epimerase showed a major protein band running near that location. Pyridoxal phosphate antagonists did not uniformly inhibit epimerase activity, but the inhibitory effect of hydroxylamine could be partially reversed by pyridoxal phosphate.
mesh_terms: D000591:Amino Acid Isomerases / Q000302:isolation & purification* / Q000378:metabolism; D001426:Bacterial Proteins; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D003067:Coenzymes / Q000378:metabolism; D003500:Cyclization; D004591:Electrophoresis, Polyacrylamide Gel; D019811:Hydroxylamine; D006898:Hydroxylamines / Q000494:pharmacology; D008970:Molecular Weight; D009842:Oligopeptides / Q000378:metabolism; D010406:Penicillins / Q000096:biosynthesis; D011732:Pyridoxal Phosphate / Q000494:pharmacology; D013302:Streptomyces / Q000201:enzymology*; D013696:Temperature
chemical_list: D001426:Bacterial Proteins; D003067:Coenzymes; D006898:Hydroxylamines; D009842:Oligopeptides; D010406:Penicillins; C011638:5-(2-aminoadipyl)cysteinylvaline; D019811:Hydroxylamine; D011732:Pyridoxal Phosphate; D000591:Amino Acid Isomerases; C040886:isopenicillin N epimerase; C006414:penicillin N