Pruess D L, Johnson M J
J Bacteriol. 1967 Nov;94(5):1502-8. doi: 10.1128/jb.94.5.1502-1508.1967.
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.
在产黄青霉的无细胞提取物中观察到青霉素与6-氨基青霉烷酸（6-APA）之间的（35）S同位素交换。活性需要含巯基的化合物。二硫苏糖醇、二硫赤藓糖醇、巯基乙醇和谷胱甘肽可作为激活剂。通过在pH 6下吸附于磷酸钙凝胶并在pH 8下洗脱，酰基转移酶被纯化了三倍。部分纯化的酶在pH 8时表现出最大活性。该酶在pH 8下于25℃至少稳定30分钟。在酶制剂中未发现除含巯基化合物以外的可解离抑制剂或激活剂。在6-APA浓度为5 mM时，测得青霉素G的表观米氏常数为1.5±0.5 mM。该酶似乎不具有青霉素酰胺酶活性。交换机制可能涉及酰基酶中间体。青霉素V、G、K、X和二氢F与（35）S-6-APA表现出同位素交换。青霉素N、甲基青霉素和苯基青霉素未表现出交换。在发酵过程中的不同时间测量了产黄青霉51-20F3中酰基转移酶的水平。活性水平在40至55小时之间增加了三倍，直到约90小时一直保持较高水平。
