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. 1995 Aug;177(15):4327–4332. doi: 10.1128/jb.177.15.4327-4332.1995

Substrate requirements for ErmC' methyltransferase activity.

P Zhong 1, S D Pratt 1, R P Edalji 1, K A Walter 1, T F Holzman 1, A G Shivakumar 1, L Katz 1
PMCID: PMC177180  PMID: 7543473

Abstract

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.

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Selected References

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