Protein-induced folding of a group I intron in cytochrome b pre-mRNA.
Some group I introns have been shown to be self-splicing in vitro, but perhaps all require proteins for splicing in vivo. Sequence differences affect the stability of secondary structures and may explain why some group I introns function efficiently without protein cofactors while others require them. The terminal intron of the cytochrome b pre-mRNA from yeast mitochondria needs a nucleus-encoded protein for splicing, even though it splices autocatalytically in high salt in vitro. This system has the advantage that the protein is specific for this intron, and yet the structure of the catalytically active RNA can be studied in its absence. We have modified the intron by chemical and enzymatic treatment in the presence and absence of the protein to determine the impact of the protein on the secondary and tertiary structures of the intron. We found protein-induced formation of secondary and tertiary structures within the intron, and the same structures also form in high salt autocatalytic conditions. We have also studied UV cross-links to determine those bases of the intron that interact directly with the protein and found that the protein contacts the intron most intimately at the structures denoted P1, L2, P4, and P6a.
Shaw
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LC
Department of Molecular Genetics and Microbiology, University of Florida, College of Medicine, Gainesville 32610-0266, USA.
Lewin
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AS
D001483:Base Sequence; D001665:Binding Sites; D002467:Cell Nucleus / Q000378:metabolism; D003573:Cytochrome b Group / Q000096:biosynthesis*; D005609:Free Radicals; D015870:Gene Expression; D015696:Gene Products, tat / Q000378:metabolism; D006678:HIV / Q000378:metabolism; D017665:Hydroxyl Radical; D007438:Introns; D008928:Mitochondria / Q000378:metabolism; D008961:Models, Structural; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009690:Nucleic Acid Conformation; D012322:RNA Precursors / Q000096:biosynthesis / Q000737:chemistry* / Q000378:metabolism*; D012326:RNA Splicing; D012331:RNA, Fungal / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism; D015183:Restriction Mapping; D012259:Ribonuclease, Pancreatic; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D014158:Transcription, Genetic; D054322:tat Gene Products, Human Immunodeficiency Virus
D003573:Cytochrome b Group; D005609:Free Radicals; D015696:Gene Products, tat; D012322:RNA Precursors; D012331:RNA, Fungal; D054322:tat Gene Products, Human Immunodeficiency Virus; D017665:Hydroxyl Radical; D012259:Ribonuclease, Pancreatic