Conformational transitioons of polypeptide chain elongation factor Tu. II. Further studies by electron spin resonance.
The conformational transitions of polypeptide chain elongation factor Tu (EF-Tu) associated with the ligand change from GDP to GTP and also with the displacement of GDP by elongation factor Ts (EF-Ts) have been investigated using the spin-labeling technique. Of the two reactive sulfhydryl groups in EF-Tu, the one essential for interaction with aminoacyl-tRNA was selectively labeled with various kinds of iodoacetamide or maleimide spin-labeling reagents. The electron spin resonance (ESR) spectra of EF-Tu-GDP labeled with these reagents generally consisted of two components, one narrow and one broad, corresponding to labels relatively weakly and strongly immobilized, respectively. The degree of immobilization and the ratio of the narrow to the broad components were different for each kind of label used. The spectra of spin-labeled EF-Tu-GDP changed markedly when its GDP moiety was replaced by GTP through incubation with phosphoenolpyruvate and pyruvate kinase [EC 2.7.1.40], the broad component increasing at the expense of the narrow component. The reversible nature of the conformational change was confirmed with EF-Tu labeled with a maleimide reagent. The GTP-induced spectral change was reversed upon conversion of labeled EF-Tu-GTP to EF-Tu-GDP by addition of excess GDP. A similar type of spectral change was also observed when spin-labeled EF-Tu-GDP was incubated with EF-Ts to form labeled EF-Tu-EF-Ts complex. The extent of the spectral change induced by EF-Ts was even greater than that induced by GTP. These results, together with those obtained by studies with hydrophobic and fluorescent probes (Arai, Arai, Kawakita, & Kaziro (1975) J. Biochem. 77, 1095-1106) indicate that a reversible conformational change is induced in EF-Tu near the sulfhydryl group that is essential for interaction with aminoacyl-tRNA.
D001665:Binding Sites; D004578:Electron Spin Resonance Spectroscopy; D006150:Guanine Nucleotides; D006160:Guanosine Triphosphate; D007460:Iodoacetamide / Q000031:analogs & derivatives; D008301:Maleimides; D010445:Peptide Elongation Factors; D011487:Protein Conformation; D013113:Spin Labels; D013438:Sulfhydryl Compounds
D006150:Guanine Nucleotides; D008301:Maleimides; D010445:Peptide Elongation Factors; D013113:Spin Labels; D013438:Sulfhydryl Compounds; D006160:Guanosine Triphosphate; D007460:Iodoacetamide