Purification, crystallization and preliminary X-ray crystallographic analysis of 23S RNA m(2)G2445 methyltransferase RlmL from Escherichia coli

Acta Crystallogr Sect F Struct Biol Cryst Commun. 2010 Nov 1;66(Pt 11):1484-6. doi: 10.1107/S1744309110035074. Epub 2010 Oct 28.

Abstract

The RlmL (YcbY) protein in Escherichia coli is an rRNA methyltransferase that is specific for m(2)G2445 modification of 23S RNA. The rlmL gene was cloned into the expression vector pET28a and expressed in the host E. coli strain BL21 (DE3). Recombinant protein with a six-histidine tag was purified by Ni(2+)-affinity chromatography followed by gel filtration. Crystals were grown using the hanging-drop vapour-diffusion method and a detergent was used as an additive to improve diffraction quality. The final crystals diffracted to 2.2 Å resolution. The crystals belonged to space group P2(1), with unit-cell parameters a = 73.6, b = 140.8, c = 102.9 Å, β = 102.3°. The crystal has a most probable solvent content of 62.8% with two molecules in the asymmetric unit.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Crystallization
  • Crystallography, X-Ray
  • Escherichia coli / enzymology*
  • Escherichia coli Proteins / chemistry*
  • Escherichia coli Proteins / isolation & purification
  • Methyltransferases / chemistry*
  • Methyltransferases / isolation & purification

Substances

  • Escherichia coli Proteins
  • Methyltransferases
  • m(2)G rRNA methyltransferase