title: The role of Glu259 in Escherichia coli elongation factor Tu in ternary complex formation.
abstract: Determination of the crystal structure of the ternary complex formed between elongation factor Tu:GTP and aminoacylated tRNA revealed three regions of interaction between elongation factor Tu and tRNA. The structure indicates that the conserved glutamic acid at position 271 in Thermus aquaticus EF-Tu could be involved in the binding of the 3' CCA-Phe end of the aminoacylated tRNA. Therefore, the corresponding residue, Glu259, of Escherichia coli EF-Tu was mutated into alanine, aspartic acid, glutamine and tyrosine, in order to substantiate the crystallographic structural evidence and to obtain further knowledge of the importance of this residue. All of the mutated proteins showed nucleotide binding properties similar to the wild type. In addition the GTPase activities were similar to the wild type. The mutation of Glu259 to either alanine or aspartic acid resulted in a reduced strength of interaction with tRNA, while mutation to tyrosine abolished completely the interaction with tRNA. Finally, mutation to glutamine resulted in an elongation factor Tu variant behaving like the wild type. In conclusion, the environment around the site binding the CCA-Phe end of the tRNA is very restricted spatially and chemically so that only a residue with almost the same size and chemical properties as glutamic acid fulfils the requirements with regard to size, salt bridge-formation potential and maintenance of the backbone conformation at the 259 position.
authors.affiliation: Institute of Molecular and Structural Biology, Aarhus University, Denmark.
authors.forename: G N
authors.initials: GN
authors.lastname: Pedersen
authors.forename: T
authors.initials: T
authors.lastname: Rattenborg
authors.forename: C R
authors.initials: CR
authors.lastname: Knudsen
authors.forename: B F
authors.initials: BF
authors.lastname: Clark
mesh_terms: D000595:Amino Acid Sequence; D017124:Conserved Sequence; D004355:Drug Stability; D004926:Escherichia coli / Q000737:chemistry* / Q000235:genetics; D020963:GTP Phosphohydrolase-Linked Elongation Factors / Q000378:metabolism; D018698:Glutamic Acid; D006153:Guanosine Diphosphate / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000378:metabolism; D006358:Hot Temperature; D006860:Hydrogen Bonding; D016297:Mutagenesis, Site-Directed; D010444:Peptide Elongation Factor Tu / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011487:Protein Conformation; D012360:RNA, Transfer, Phe / Q000378:metabolism*; D013329:Structure-Activity Relationship
chemical_list: D012360:RNA, Transfer, Phe; D006153:Guanosine Diphosphate; D018698:Glutamic Acid; D006160:Guanosine Triphosphate; D020963:GTP Phosphohydrolase-Linked Elongation Factors; D010444:Peptide Elongation Factor Tu