The effect of long-range loop-loop interactions on folding of the Tetrahymena self-splicing RNA.
Bas?e-pairing between the terminal loops of helices P2.1 and P9.1a (P13) and P2 and P5c (P14) stabilize the folded structure of the Tetrahymena group I intron. Using native gel electrophoresis to analyze the folding kinetics of a natural pre-RNA containing the Tetrahymena intron, we show that P13 and P14 are the only native loop-loop interactions among six possible combinations. Other base-pairing interactions of the loop sequences stabilize misfolded and inactive pre-RNAs. Mismatches in P13 or P14 raised the midpoints and decreased the cooperativity of the Mg(2+)-dependent eqXuilibrium folding transitions. Although some mutations in P13 resulted in slightly higher folding rates, others led to slower folding compared to the wild-type, suggesting that P13 promotes formation of P3 and P7. In contrast, mismatches in P14 increased the rate of folding, suggesting that base-pairing between P5c and P2 stabilizes intermediates in which the catalytic core is misfolded. Although the peripheral helices stabilize the native structure of the catalytic core, our results show that formation of long-range interactions, and competition between correct and incorrect loop-loop base-pairs, decrease the rate at which the active pre-RNA structure is assembled.
Pan
J
J
College Park, University of Maryland, College Park, MD 20742-2021, USA.
Woodson
S A
SA
D000818:Animals; D001483:Base Sequence; D001667:Binding, Competitive; D007438:Introns; D007700:Kinetics; D008274:Magnesium / Q000378:metabolism; D008958:Models, Molecular; D009154:Mutation; D009690:Nucleic Acid Conformation; D009694:Nucleic Acid Precursors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012326:RNA Splicing; D020871:RNA Stability; D016337:RNA, Catalytic / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D016053:RNA, Protozoan / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D013768:Tetrahymena / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
D009694:Nucleic Acid Precursors; D016337:RNA, Catalytic; D016053:RNA, Protozoan; D008274:Magnesium