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Mapping the fMet‐tRNA f Met binding site of initiation factor IF2
Author(s) -
Guenneugues Marc,
Caserta Enrico,
Brandi Letizia,
Spurio Roberto,
Meunier Sylvie,
Pon Cynthia L.,
Boelens Rolf,
Gualerzi Claudio O.
Publication year - 2000
Publication title -
the embo journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 7.484
H-Index - 392
eISSN - 1460-2075
pISSN - 0261-4189
DOI - 10.1093/emboj/19.19.5233
Subject(s) - biology , transfer rna , eukaryotic translation , initiation factor , binding site , stereochemistry , site directed mutagenesis , directed mutagenesis , acceptor , biochemistry , crystallography , translation (biology) , chemistry , gene , rna , messenger rna , physics , mutant , condensed matter physics
The interaction between fMet‐tRNA f Met and Bacillus stearothermophilus translation initiation factor IF2 has been characterized. We demonstrate that essentially all thermodynamic determinants governing the stability and the specificity of this interaction are localized within the acceptor hexanucleotide fMet‐3′ACCAAC of the initiator tRNA and a fairly small area at the surface of the β‐barrel structure of the 90‐amino acid C‐terminal domain of IF2 (IF2 C‐2). A weak but specific interaction between IF2 C‐2 and formyl‐methionyl was also demonstrated. The surface of IF2 C‐2 interacting with fMet‐tRNA f Met has been mapped using two independent approaches, site‐ directed mutagenesis and NMR spectroscopy, which yielded consistent results. The binding site comprises C668 and G715 located in a groove accommodating the methionyl side‐chain, R700, in the vicinity of the formyl group, Y701 and K702 close to the acyl bond between fMet and tRNA f Met , and the surface lined with residues K702‐S660, along which the acceptor arm of the initiator tRNA spans in the direction 3′ to 5′.