The location and the significance of a cross-link between the sarcin/ricin domain of ribosomal RNA and the elongation factor-G.
During translocation peptidyl-tRNA moves from the A-site to the P-site and mRNA is displaced by three nucleotides in the 3' direction. This reaction is catalyzed by elongation factor-G (EF-G) and is associated with ribosome-dependent hydrolysis of GTP. The molecular basis of translocation is the most important unsolved problem with respect to ribosome function. A critical question, one that might provide a clue to the mechanism of translocation, is the precise identity of the contacts between EF-G and ribosome components. To make the identification, a covalent bond was formed, by ultraviolet irradiation, between EF-G and a sarcin/ricin domain (SRD) oligoribonucleotide containing 5-iodouridine. The cross-link was established, by mass spectroscopy and by Edman degradation, to be between a tryptophan at position 127 in the G domain in EF-G and either one of two 5-iodouridine nucleotides in the sequence UAG2655U in the SRD. G2655 is a critical identity element for the recognition of the factor's ribosomal binding site. The site of the cross-link provides the first direct evidence that the SRD is in close proximity to the EF-G catalytic center. The proximity suggests that the SRD RNA has a role in the activation of GTP hydrolysis that leads to a transition in the conformation of the factor and to its release from the ribosome.
Department of Biochemistry and Molecular Biology, The University of Chicago, 920 East 58th Street, Chicago, IL 60637, USA.
Yuen-Ling
YL
Chan
Carl C
CC
Correll
Ira G
IG
Wool
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D003432:Cross-Linking Reagents; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D046911:Macromolecular Substances; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D020653:Peptide Elongation Factor G / Q000737:chemistry* / Q000378:metabolism*; D012329:RNA, Bacterial / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012335:RNA, Ribosomal / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D012276:Ricin / Q000737:chemistry* / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D016963:Thermus thermophilus / Q000235:genetics / Q000378:metabolism; D014466:Ultraviolet Rays
D001426:Bacterial Proteins; D003432:Cross-Linking Reagents; D029968:Escherichia coli Proteins; D046911:Macromolecular Substances; D020653:Peptide Elongation Factor G; D012329:RNA, Bacterial; D012335:RNA, Ribosomal; D012276:Ricin