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Proteins. 2005 May 15;59(3):482-8.

Sequence-structure-function relationships of a tRNA (m7G46) methyltransferase studied by homology modeling and site-directed mutagenesis.

Author information

1
Laboratory of Bioinformatics and Protein Engineering, International Institute of Molecular and Cell Biology, Warsaw, Poland.

Abstract

The Escherichia coli TrmB protein and its Saccharomyces cerevisiae ortholog Trm8p catalyze the S-adenosyl-L-methionine-dependent formation of 7-methylguanosine at position 46 (m7G46) in tRNA. To learn more about the sequence-structure-function relationships of these enzymes we carried out a thorough bioinformatics analysis of the tRNA:m7G methyltransferase (MTase) family to predict sequence regions and individual amino acid residues that may be important for the interactions between the MTase and the tRNA substrate, in particular the target guanosine 46. We used site-directed mutagenesis to construct a series of alanine substitutions and tested the activity of the mutants to elucidate the catalytic and tRNA-recognition mechanism of TrmB. The functional analysis of the mutants, together with the homology model of the TrmB structure and the results of the phylogenetic analysis, revealed the crucial residues for the formation of the substrate-binding site and the catalytic center in tRNA:m7G MTases.

PMID:
15789416
DOI:
10.1002/prot.20454
[Indexed for MEDLINE]

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