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Mutational analysis of Glu 272 in elongation factor 1A of E. coli
Author(s) -
Mansilla F.,
Knudsen C.R.,
Clark B.F.C.
Publication year - 1998
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/s0014-5793(98)00646-2
Subject(s) - ef tu , elongation factor , escherichia coli , arginine , amino acid , aminoacyl trna , biochemistry , salt bridge , stereochemistry , chemistry , transfer rna , thermus thermophilus , biology , ribosome , mutant , gene , rna
In our previous work (Mansilla et al. (1997) Protein Eng. 10, 927–934) we showed that Arg 7 of Escherichia coli elongation factor Tu (EF1A) plays an essential role in aminoacyl‐tRNA (aa‐tRNA) binding. Substitution of Arg 7 by Ala or Glu lost this activity. We proposed that Arg 7 forms a salt bridge with the charged conserved amino acid Glu 272 (Asp 284 in Thermus aquaticus ) thereby binding the N‐terminal region of the protein to domain 2 and thus completing the conformational rearrangement needed for binding aa‐tRNA. In this work we have mutated Glu 272 to arginine, either alone (Glu 272 Arg), or in combination with one of the above mentioned mutations (Arg 7 Glu/Glu 272 Arg) in order to test this hypothesis. Our results show that, in confirmation of our thesis based on structural knowledge, the substitution of Glu 272 (Asp 284 ) decreases the ability of EF1A:GTP to bind aa‐tRNA.