Recognition of the initiator tRNA by the Pseudomonas aeruginosa methionyl-tRNA formyltransferase: importance of the base-base mismatch at the end of the acceptor stem.
Formylation of the initiator methionyl-tRNA (Met-tRNAfMet) in eubacteria is catalyzed by methionyl-tRNA formyltransferase (MTF). Features of the Escherichia coli tRNAfMet that are important for formylation are the base-base mismatch between nucleotides 1 and 72, and the second and third base pairs of the acceptor stem. The base-base mismatch is the most crucial formylation determinant in the E. coli tRNAfMet. However, it is not known whether this feature is also important for formylation of other eubacterial tRNAfMet. We cloned the Pseudomonas aeruginosa MTF gene by complementation of an E. coli MTF mutant strain with a genomic library, and investigated the catalytic properties and substrate specificity of the enzyme. The results show that the P. aeruginosa and E. coli enzymes have comparable affinities for the tRNAfMet and N10-formyltetrahydrofolate (fTHF) substrates. Overproduction of the P. aeruginosa MTF rescued the initiator activity of an E. coli formylation-defective tRNAfMet with a base pair between nucleotides 1 and 72, indicating that the base-base mismatch is utilized by the P. aeruginosa MTF for recognition of the tRNAfMet. Therefore, this feature may be used by MTFs from other eubacteria to distinguish the initiator from elongator tRNAs.
Newton
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Guelph-Waterloo Center for Graduate Work in Chemistry and Biochemistry, Department of Chemistry and Biochemistry, University of Guelph, Ont., Canada.
Niemkiewicz
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Mangroo
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D000595:Amino Acid Sequence; D020137:Base Pair Mismatch; D001483:Base Sequence; D002384:Catalysis; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D005575:Formyltetrahydrofolates / Q000378:metabolism; D005798:Genes, Bacterial; D005816:Genetic Complementation Test; D019877:Hydroxymethyl and Formyl Transferases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D010442:Peptide Chain Initiation, Translational; D011550:Pseudomonas aeruginosa / Q000201:enzymology / Q000235:genetics* / Q000378:metabolism*; D012358:RNA, Transfer, Met / Q000378:metabolism*; D013379:Substrate Specificity
D005575:Formyltetrahydrofolates; D012358:RNA, Transfer, Met; C024988:tRNA, formylmethionine-; D019877:Hydroxymethyl and Formyl Transferases; C021561:methionyl-tRNA formyltransferase