title: Purification, crystallization and preliminary X-ray diffraction analysis of the 23S rRNA methyltransferase RlmJ from Escherichia coli.
abstract: Methyltransferase RlmJ uses the cofactor S-adenosylmethionine to methylate the exocyclic nitrogen N6 of nucleotide A2030 in 23S rRNA during ribosome assembly in Escherichia coli. RlmJ with a C-terminal hexahistidine tag was overexpressed in E. coli and purified as a monomer using Ni(2+)-affinity and size-exclusion chromatography. The recombinant RlmJ was crystallized using the sitting-drop vapour-diffusion method and a full data set was collected to 1.85 Å resolution from a single apo crystal. The crystals belonged to space group P2(1), with unit-cell parameters a = 46.9, b = 77.8, c = 82.5 Å, β = 104°. Data analysis suggested two molecules per asymmetric unit and a Matthews coefficient of 2.20 Å(3) Da(-1).
authors.affiliation: Department of Cell and Molecular Biology, Uppsala University, PO Box 596, 751 24 Uppsala, Sweden.
authors.forename: Avinash S
authors.initials: AS
authors.lastname: Punekar
authors.forename: Maria
authors.initials: M
authors.lastname: Selmer
mesh_terms: D000595:Amino Acid Sequence; D018360:Crystallography, X-Ray; D004926:Escherichia coli / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D015870:Gene Expression; D008780:Methyltransferases / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D008969:Molecular Sequence Data; D054730:Protein Interaction Domains and Motifs; D012338:RNA, Ribosomal, 23S / Q000737:chemistry*; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D012270:Ribosomes / Q000235:genetics / Q000378:metabolism; D012436:S-Adenosylmethionine / Q000737:chemistry*
chemical_list: D029968:Escherichia coli Proteins; D012338:RNA, Ribosomal, 23S; D011993:Recombinant Fusion Proteins; D012436:S-Adenosylmethionine; D008780:Methyltransferases; C585479:RlmJ protein, E coli
keywords: 23S rRNA; Escherichia coli; RlmJ; S-adenosylmethionine; m6A2030; methyltransferases; ribosome assembly