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.
Chan
Yuen-Ling
YL
Department of Biochemistry and Molecular Biology, The University of Chicago, 920 East 58th Street, Chicago, IL 60637, USA.
Correll
Carl C
CC
Wool
Ira G
IG
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