Mechanistic insights on the folding of a large ribozyme during transcription.
RNA folding during transcription resembles folding in a cellular environment. We previously investigated the folding of a large ribozyme derived from a bacterial RNase P RNA during its transcription by the Escherichia coli RNA polymerase and the effect of the elongation factor NusA. We found that transcriptional pausing at a specific site induced by NusA significantly altered the folding pathway. In this work, we compare folding during transcription by the E. coli RNA polymerase of circularly permuted variants and site-specific mutants of the RNase P ribozyme to elucidate the molecular mechanism of transcriptional pausing and RNA folding. The effect of NusA-induced pausing depends on the order of RNA synthesis and only affects local folding of the RNA. Pausing likely prevents a misfolded structure between the 5' strand of a helix and its adjacent junction located in the specificity domain and a region known to bind single-stranded RNA located in the catalytic domain. These results lead to a structural model on how transcriptional pausing affects folding of RNase P RNA. Structural rearrangements of a nascent RNA transcript enhanced by transcriptional pausing may be a general feature of RNA folding during transcription.
Wong
Terrence
T
Department of Biochemistry and Molecular Biology, University of Chicago, Chicago, Illinois 60637, USA.
Sosnick
Tobin R
TR
Pan
Tao
T
D001483:Base Sequence; D002384:Catalysis; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000235:genetics; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D009690:Nucleic Acid Conformation; D009693:Nucleic Acid Hybridization; D010445:Peptide Elongation Factors / Q000737:chemistry; D016337:RNA, Catalytic / Q000737:chemistry* / Q000235:genetics*; D043262:Ribonuclease P / Q000737:chemistry / Q000235:genetics; D014157:Transcription Factors / Q000737:chemistry; D014158:Transcription, Genetic; D035602:Transcriptional Elongation Factors
D029968:Escherichia coli Proteins; D010445:Peptide Elongation Factors; D016337:RNA, Catalytic; D014157:Transcription Factors; D035602:Transcriptional Elongation Factors; C068727:nusA protein, E coli; D043262:Ribonuclease P