期刊
JOURNAL OF MOLECULAR BIOLOGY
卷 315, 期 3, 页码 297-310出版社
ACADEMIC PRESS LTD ELSEVIER SCIENCE LTD
DOI: 10.1006/jmbi.2001.5233
关键词
RNA folding; footprinting; kinetics; group II ribozyme; magnesium titration
资金
- NCRR NIH HHS [P41-RR 01633] Funding Source: Medline
- NIDDK NIH HHS [DK 07328] Funding Source: Medline
- NIGMS NIH HHS [R01 GM 50313, T32 GM 08281] Funding Source: Medline
Group II introns are large catalytic RNA molecules that fold into compact structures essential for the catalysis of splicing and intron mobility reactions. Despite a growing body of information on the folded state of group II introns at equilibrium, there is currently no information on the folding pathway and little information on the ionic requirements for folding. Folding isotherms were determined by hydroxyl radical footprinting for the 32 individual protections that are distributed throughout a group II intron ribozyme derived from intron ai5gamma. The isotherms span a similar range of Mg2+ concentrations and share a similar index of cooperativity. Time-resolved hydroxyl radical footprinting studies show that all regions of the ribozyme fold slowly and with remarkable synchrony into a single catalytically active structure at a rate comparable to those of other ribozymes studied thus far. The rate constants for the formation of tertiary contacts and recovery of catalytic activity are identical within experimental error. Catalytic activity analyses in the presence of urea provide no evidence that the slow folding of the ai5gamma intron is attributable to the presence of unproductive kinetic traps along the folding pathway. Taken together, the data suggest that the rate-limiting step for folding of group II intron ai5gamma occurs early along the reaction pathway. We propose that this behavior resembles protein folding that is limited in rate by high contact order, or the need to form key tertiary interactions from partners that are located far apart in the primary or secondary structure. (C) 2002 Academic Press.
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