PMID:15158725

Citation	Hunter, SE and Spremulli, LL (2004) Effects of mutagenesis of residue 221 on the properties of bacterial and mitochondrial elongation factor EF-Tu. Biochim. Biophys. Acta 1699:173-82
Abstract	During protein biosynthesis, elongation factor Tu (EF-Tu) delivers aminoacyl-tRNA (aa-tRNA) to the A-site of ribosomes. This factor is highly conserved throughout evolution. However, several key residues differ between bacterial and mammalian mitochondrial EF-Tu (EF-Tu(mt)). One such residue is Ser221 (Escherichia coli numbering). This residue is conserved as a Ser or Thr in the bacterial factors but is present as Pro269 in EF-Tu(mt). Pro269 reorients the loop containing this residue and shifts the adjoining beta-strand in EF-Tu(mt) compared to that of E. coli EF-Tu potentially altering the binding pocket for the acceptor stem of the aa-tRNA. Pro269 was mutated to a serine residue (P269S) in EF-Tu(mt). For comparison, the complementary mutation was created at Ser221 in E. coli EF-Tu (S221P). The E. coli EF-Tu S221P variant is poorly expressed in E. coli and the majority of the molecules fail to fold into an active conformation. In contrast, EF-Tu(mt) P269S is expressed to a high level in E. coli. When corrected for the percentage of active molecules, both variants function as effectively as their respective wild-type factors in ternary complex formation using E. coli Phe-tRNA(Phe) and Cys-tRNA(Cys). They are also active in A-site binding and in vitro translation assays with E. coli Phe-tRNA(Phe). In addition, both variants are as active as their respective wild-type factors in ternary complex formation, A-site binding and in vitro translation assays using mitochondrial Phe-tRNA(Phe).
Links	PubMed Online version:10.1016/j.bbapap.2004.02.015
Keywords	Amino Acid Sequence; Escherichia coli/chemistry; Escherichia coli/genetics; Escherichia coli/metabolism; Guanine Nucleotides/metabolism; Mitochondria/chemistry; Mitochondria/metabolism; Molecular Sequence Data; Mutagenesis; Peptide Elongation Factor Tu/chemistry; Peptide Elongation Factor Tu/genetics; Peptide Elongation Factor Tu/metabolism; Protein Biosynthesis; Protein Conformation; RNA, Transfer, Cys/chemistry; RNA, Transfer, Phe/chemistry; Ribosomes/metabolism; Sequence Homology, Amino Acid
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