mRNA binding track in the human 80S ribosome for mRNA analogues randomly substituted with 4-thiouridine residues.
The interaction between mRNA and 18S rRNA in human 80S ribosomes has been studied using synthetic mRNA analogues randomly substituted with 4-thiouridine, which can be photoactivated for cross-linking. Two mRNA analogues with different sequences have been used for complex formation with ribosomes without or with the presence of a cognate tRNA. Cross-linked 18S rRNA nucleotides were identified by reverse transcription analysis. The base U630 in 18S rRNA was the main target of cross-linking for both of the mRNA analogues studied, and three minor sites of cross-linking, A1060, U1046, and U966, were also identified. Thus, in the case of human 80S ribosomes, the set of nucleotide residues cross-linked to the mRNA analogues is significantly smaller than the twelve sites seen for Escherichia coli with these same two mRNA analogues [Bhangu, R., & Wollenzien, P. (1992) Biochemistry 31, 5937-5944]. The residue U630 is within a highly conserved region corresponding to the 530 loop region of eubacterial 16S rRNA; the cross-link to this site indicates that it plays a key role in interacting with mRNA on 80S ribosomes independently of the presence of a cognate tRNA at the P site.
Graifer
D M
DM
Department of Biochemistry, North Carolina State University, Raleigh 27695.
Juzumiene
D I
DI
Karpova
G G
GG
Wollenzien
P
P
D001483:Base Sequence; D001665:Binding Sites; D003432:Cross-Linking Reagents; D004587:Electrophoresis, Agar Gel; D006801:Humans; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D012333:RNA, Messenger / Q000737:chemistry / Q000378:metabolism*; D012337:RNA, Ribosomal, 18S / Q000378:metabolism*; D012360:RNA, Transfer, Phe / Q000378:metabolism; D012364:RNA, Transfer, Trp / Q000378:metabolism; D012194:RNA-Directed DNA Polymerase / Q000378:metabolism; D012270:Ribosomes / Q000378:metabolism*; D013891:Thiouridine; D014466:Ultraviolet Rays
D003432:Cross-Linking Reagents; D012333:RNA, Messenger; D012337:RNA, Ribosomal, 18S; D012360:RNA, Transfer, Phe; D012364:RNA, Transfer, Trp; D013891:Thiouridine; D012194:RNA-Directed DNA Polymerase