The methylation of one specific guanosine in a pre-tRNA prevents cleavage by RNase P and by the catalytic M1 RNA.
Several modified nucleosides were introduced during in vitro RNA synthesis into a pre-tRNA(Ser). The pre-tRNAs were used as substrates for RNase P enzymes. No effects were observed with biotin-8-ATP or [alpha-S]-GPT, whereas with m7GTP, the cleavage reaction was completely inhibited. Analysis of pre-tRNAs which contained m7G at various positions has revealed a single base at the 5'-end of the acceptor stem where this modification absolutely prevents cleavage by catalytic M1 RNA, eukaryotic and prokaryotic RNase P holoenzymes. These results suggest that a critical contact must be made between pre-tRNA substrate and enzyme/ribozyme or that the approach of the potential cleaving agent (a positive magnesium ion) is made impossible by the positive charge at N-7 of the guanosine. In addition, we have shown that a pre-tRNA containing only m7G's can still form a complex with M1 RNA in a gel retardation assay.
Kahle
D
D
Institute for General Microbiology, Christian-Albrechts-Universität, Kiel, FRG.
Wehmeyer
U
U
Char
S
S
Krupp
G
G
D001483:Base Sequence; D004722:Endoribonucleases / Q000378:metabolism*; D005800:Genes, Fungal; D006151:Guanosine; D008745:Methylation; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D010957:Plasmids; D012322:RNA Precursors / Q000235:genetics; D016337:RNA, Catalytic; D012335:RNA, Ribosomal / Q000378:metabolism*; D012343:RNA, Transfer / Q000235:genetics; D043262:Ribonuclease P; D012568:Schizosaccharomyces / Q000235:genetics; D013379:Substrate Specificity
D012322:RNA Precursors; D016337:RNA, Catalytic; D012335:RNA, Ribosomal; D006151:Guanosine; D012343:RNA, Transfer; D004722:Endoribonucleases; D043262:Ribonuclease P