Bauer N J, Kreuzman A J, Dotzlaf J E, Yeh W K
Biochemical Development Division, Eli Lilly and Company, Indianapolis, Indiana 46285.
J Biol Chem. 1988 Oct 25;263(30):15619-25.
S-Adenosyl-L-methionine:macrocin O-methyltransferase catalyzes conversion of macrocin to tylosin, the terminal and main rate-limiting step of tylosin biosynthesis in Streptomyces fradiae. The O-methyltransferase was stabilized in vitro and purified to electrophoretic homogeneity. The purified enzyme had a molecular weight of 65,000 and consisted of two identical subunits of 32,000 with an isoelectric point of 4.5. The enzyme required Mg2+, Mn2+, or Co2+ for maximal activity and was catalytically optimal at pH 7.5-8.0 and 31 degrees C. The O-methyltransferase catalyzed the conversion of macrocin to tylosin at a stoichiometric ratio of 1:1. The enzyme also mediated conversion of lactenocin----desmycosin. The corresponding Vmax/Km ratios for the two analogous conversions were similar, and both enzymic conversions were susceptible to extensive competitive and noncompetitive inhibitions by macrolide metabolites. Steady-state kinetic studies for initial velocity, substrate analogue, and product inhibitions have allowed formulation of Ordered Bi Bi as the reaction mechanism for macrocin O-methyltransferase.
S-腺苷-L-甲硫氨酸：大环菌素O-甲基转移酶催化大环菌素转化为泰乐菌素，这是弗氏链霉菌中泰乐菌素生物合成的末端且主要的限速步骤。该O-甲基转移酶在体外得到稳定并纯化至电泳纯。纯化后的酶分子量为65,000，由两个相同的32,000亚基组成，等电点为4.5。该酶需要Mg2+、Mn2+或Co2+以达到最大活性，在pH 7.5 - 8.0和31℃时催化效果最佳。O-甲基转移酶以1:1的化学计量比催化大环菌素转化为泰乐菌素。该酶还介导了乳内霉素向去碳霉糖泰乐菌素的转化。这两种类似转化的相应Vmax/Km比值相似，并且两种酶促转化都受到大环内酯类代谢物广泛的竞争性和非竞争性抑制。对初始速度、底物类似物和产物抑制的稳态动力学研究使得有序双底物双产物机制被确定为大环菌素O-甲基转移酶的反应机制。
