Cloning and characterization of tRNA (m1A58) methyltransferase (TrmI) from Thermus thermophilus HB27, a protein required for cell growth at extreme temperatures.
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.
Droogmans
Louis
L
Laboratoire de Microbiologie, Université Libre de Bruxelles, Institut de Recherches Microbiologiques Jean-Marie Wiame, Avenue E. Gryson 1, B-1070 Bruxelles, Belgium.
Roovers
Martine
M
Bujnicki
Janusz M
JM
Tricot
Catherine
C
Hartsch
Thomas
T
Stalon
Victor
V
Grosjean
Henri
H
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
D012349:RNA, Transfer, Asp; D012359:tRNA Methyltransferases; C023194:tRNA (adenine(58)-N(1))-methyltransferase