Structure of M1 RNA as determined by psoralen cross-linking.
The RNA moiety of ribonuclease P from Escherichia coli (M1 RNA) has been photoreacted with 4'-(hydroxymethyl)-4,5',8-trimethylpsoralen (HMT) and long-wave UV light (320-380 nm) in a buffer containing 60 mM Mg2+, where the RNA moiety acts as a true catalyst of tRNA processing. Limited specific digestion and two-dimensional gel electrophoresis yield fragments cross-linked by HMT. By photoreversal of the isolated cross-linked fragments and enzymatic sequencing of the fragments, the positions of the cross-links have been elucidated. This method allows us to locate the cross-link to +/- 15 nucleotides. Further assignments of the exact locations of the cross-links have been made on the basis of the known photoreactivity of the psoralen with different bases. Nine unique cross-links have been isolated in the M1 RNA including four long-range interactions. The short-range interactions are discussed here in detail.
D001483:Base Sequence; D003432:Cross-Linking Reagents; D004591:Electrophoresis, Polyacrylamide Gel; D004722:Endoribonucleases / Q000302:isolation & purification; D029968:Escherichia coli Proteins; D011564:Furocoumarins; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D012329:RNA, Bacterial; D043262:Ribonuclease P; D014307:Trioxsalen / Q000031:analogs & derivatives
D003432:Cross-Linking Reagents; D029968:Escherichia coli Proteins; D011564:Furocoumarins; D012329:RNA, Bacterial; C016507:hydroxymethyltrioxsalen; D004722:Endoribonucleases; D043262:Ribonuclease P; C470865:ribonuclease P, E coli; D014307:Trioxsalen