Affinity labeling of the GDP/GTP binding site in Thermus thermophilus elongation factor Tu.
Elongation factor Tu from Thermus thermophilus was treated successively with periodate-oxidized GDP or GTP and cyanoborohydride. Covalently modified cyanogen bromide or trypsin fragments of the protein were isolated, and the position of their modification was determined. Lysine residues 52 and 137 were heavily labeled, lysine-137 being considerably more reactive in the GTP form as compared to the GDP form of the protein. These residues are in the proximity of the GDP/GTP binding site. Lys-325 was also labeled, but to a lower extent. The part of the EF-Tu containing residue 52 is missing in crystallized EF-Tu.GDP from Escherichia coli [Jurnak, F. (1985) Science (Washington, D.C.) 230, 32-36]. These results place the part of T. thermophilus EF-Tu corresponding to the missing fragment in E. coli EF-Tu in the vicinity of the nucleotide binding site and allow its role in the interaction with aminoacyl-tRNA and elongation factor Ts to be evaluated. Cross-linking of EF-Tu.GDP by irradiation at 257 nm showed that a sequence of 10 amino acids residues which is found in the Thermus thermophilus elongation factor Tu but not in other homologous bacterial proteins is located in the vicinity of the GDP/GTP binding site.
Peter
M E
ME
Laboratorium für Biochemie, Universität Bayreuth, FRG.
Wittmann-Liebold
B
B
Sprinzl
M
M
D000345:Affinity Labels / Q000378:metabolism*; D000495:Allosteric Site; D001426:Bacterial Proteins / Q000378:metabolism*; D006150:Guanine Nucleotides / Q000378:metabolism*; D006153:Guanosine Diphosphate / Q000378:metabolism*; D006160:Guanosine Triphosphate / Q000378:metabolism*; D008958:Models, Molecular; D010444:Peptide Elongation Factor Tu / Q000378:metabolism*; D010449:Peptide Mapping; D011485:Protein Binding; D011487:Protein Conformation; D013824:Thermus / Q000378:metabolism*
D000345:Affinity Labels; D001426:Bacterial Proteins; D006150:Guanine Nucleotides; D006153:Guanosine Diphosphate; D006160:Guanosine Triphosphate; D010444:Peptide Elongation Factor Tu