title: Cloning and characterization of tRNA (m1A58) methyltransferase (TrmI) from Thermus thermophilus HB27, a protein required for cell growth at extreme temperatures.
abstract: N1-methyladenosine (m1A) is found at position 58 in the T-loop of many tRNAs. In yeast, the formation of this modified nucleoside is catalyzed by the essential tRNA (m1A58) methyltransferase, a tetrameric enzyme that is composed of two types of subunits (Gcd14p and Gcd10p). In this report we describe the cloning, expression and characterization of a Gcd14p homolog from the hyperthermophilic bacterium Thermus thermophilus. The purified recombinant enzyme behaves as a homotetramer of 150 kDa by gel filtration and catalyzes the site- specific formation of m1A at position 58 of the T-loop of tRNA in the absence of any other complementary protein. S-adenosylmethionine is used as donor of the methyl group. Thus, we propose to name the bacterial enzyme TrmI and accordingly its structural gene trmI. These results provide a key evolutionary link between the functionally characterized two-component eukaryotic enzyme and the recently described crystal structure of an uncharacterized, putative homotetrameric methyltransferase Rv2118c from Mycobacterium tuberculosis. Interest ingly, inactivation of the T.thermophilus trmI gene results in a thermosensitive phenotype (growth defect at 80 degrees C), which suggests a role of the N1-methylation of tRNA adenosine-58 in adaptation of life to extreme temperatures.
authors.affiliation: Laboratoire de Microbiologie, Université Libre de Bruxelles, Institut de Recherches Microbiologiques Jean-Marie Wiame, Avenue E. Gryson 1, B-1070 Bruxelles, Belgium.
authors.forename: Louis
authors.initials: L
authors.lastname: Droogmans
authors.forename: Martine
authors.initials: M
authors.lastname: Roovers
authors.forename: Janusz M
authors.initials: JM
authors.lastname: Bujnicki
authors.forename: Catherine
authors.initials: C
authors.lastname: Tricot
authors.forename: Thomas
authors.initials: T
authors.lastname: Hartsch
authors.forename: Victor
authors.initials: V
authors.lastname: Stalon
authors.forename: Henri
authors.initials: H
authors.lastname: Grosjean
mesh_terms: D000595:Amino Acid Sequence; D002455:Cell Division / Q000235:genetics; D002850:Chromatography, Gel; D003001:Cloning, Molecular; D019281:Dimerization; D004591:Electrophoresis, Polyacrylamide Gel; D006358:Hot Temperature; D008969:Molecular Sequence Data; D009154:Mutation; D010641:Phenotype; D012349:RNA, Transfer, Asp / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013696:Temperature; D016963:Thermus thermophilus / Q000201:enzymology / Q000235:genetics* / Q000254:growth & development; D012359:tRNA Methyltransferases / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism
chemical_list: D012349:RNA, Transfer, Asp; D012359:tRNA Methyltransferases; C023194:tRNA (adenine(58)-N(1))-methyltransferase