Kinetic analysis of the M1 RNA folding pathway.
The biological activity of large RNAs is dependent on the formation of complex folded structures that determine function. Typically the creation of such structures requires divalent magnesium and in many cases the folding process takes place over the course of several minutes. It has been proposed that the folding paths of large RNAs proceed through discrete intermediates but the nature of these intermediates is not known in most cases. Here, we describe our studies on the folding of the M1 RNA sub-unit of Escherichia coli RNase P. We performed kinetic footprinting studies of M1 RNA folding with the chemical footprinting reagent peroxynitrous acid to provide a detailed description of the folding pathway of RNase P RNA. Our results indicate that, in contrast to the Group I ribozyme, the M1 RNA folds into its catalytically active structure through the formation of two separately folded domains and that the folding of each proceeds through a discrete series of intermediates. Similar rates of folding were observed for regions believed to form the interface between the two domains. This observation is consistent with a kinetic trap which occurs by interaction of the domains during folding.
Kent
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Department of Biochemistry, University of Alberta, 4-74 Medical Sciences Building, Edmonton, Alberta, T6G 2C6, Canada.
Chaulk
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MacMillan
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D001665:Binding Sites; D002384:Catalysis; D004722:Endoribonucleases / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics*; D029968:Escherichia coli Proteins; D007700:Kinetics; D008958:Models, Molecular; D009608:Nitrous Acid / Q000378:metabolism; D009690:Nucleic Acid Conformation; D030421:Peroxynitrous Acid; D020871:RNA Stability; D012329:RNA, Bacterial / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D016337:RNA, Catalytic / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D043262:Ribonuclease P
D029968:Escherichia coli Proteins; D012329:RNA, Bacterial; D016337:RNA, Catalytic; D030421:Peroxynitrous Acid; D004722:Endoribonucleases; D043262:Ribonuclease P; C470865:ribonuclease P, E coli; D009608:Nitrous Acid