Zhong P, Pratt S D, Edalji R P, Walter K A, Holzman T F, Shivakumar A G, Katz L
Pharmaceutical Products Division, Abbott Laboratories, Abbott Park, Illinois 60064, USA.
J Bacteriol. 1995 Aug;177(15):4327-32. doi: 10.1128/jb.177.15.4327-4332.1995.
ErmC' is a methyltransferase that confers resistance to the macrolide-lincosamide-streptogramin B group of antibiotics by catalyzing the methylation of 23S rRNA at a specific adenine residue (A-2085 in Bacillus subtilis; A-2058 in Escherichia coli). The gene for ErmC' was cloned and expressed to a high level in E. coli, and the protein was purified to virtual homogeneity. Studies of substrate requirements of ErmC' have shown that a 262-nucleotide RNA fragment within domain V of B. subtilis 23S rRNA can be utilized efficiently as a substrate for methylation at A-2085. Kinetic studies of the monomethylation reaction showed that the apparent Km of this 262-nucleotide RNA oligonucleotide was 26-fold greater than the value determined for full-size and domain V 23S rRNA. In addition, the Vmax for this fragment also rose sevenfold. A model of RNA-ErmC' interaction involving multiple binding sites is proposed from the kinetic data presented.
ErmC'是一种甲基转移酶，通过催化23S rRNA在特定腺嘌呤残基（枯草芽孢杆菌中为A-2085；大肠杆菌中为A-2058）处的甲基化，赋予对大环内酯-林可酰胺-链阳霉素B组抗生素的抗性。ErmC'基因在大肠杆菌中被克隆并高水平表达，蛋白质被纯化至几乎均一。对ErmC'底物需求的研究表明，枯草芽孢杆菌23S rRNA结构域V内的一个262个核苷酸的RNA片段可有效用作A-2085甲基化的底物。单甲基化反应的动力学研究表明，这个262个核苷酸的RNA寡核苷酸的表观Km比全尺寸和结构域V 23S rRNA确定的值大26倍。此外，该片段的Vmax也提高了7倍。根据所提供的动力学数据，提出了一个涉及多个结合位点的RNA-ErmC'相互作用模型。
