An RNA chaperone activity of non-specific RNA binding proteins in hammerhead ribozyme catalysis.
We have previously shown that a protein derived from the p7 nucleocapsid (NC) protein of HIV type-1 increases kcat/Km and kcat for cleavage of a cognate substrate by a hammerhead ribozyme. Here we show directly that the increase in kcat/Km arises from catalysis of the annealing of the RNA substrate to the ribozyme and the increase in kcat arises from catalysis of dissociation of the RNA products from the ribozyme. A peptide polymer derived from the consensus sequence of the C-terminal domain of the hnRNP A1 protein (A1 CTD) provides similar enhancements. Although these effects apparently arise from non-specific interactions, not all non-specific binding interactions led to these enhancements. NC and A1 CTD exert their effects by accelerating attainment of the thermodynamically most stable species throughout the ribozyme catalytic cycle. In addition, NC protein is shown to resolve a misfolded ribozyme-RNA complex that is otherwise long lived. These in vitro results suggest that non-specific RNA binding proteins such as NC and hnRNP proteins may have a biological role as RNA chaperones that prevent misfolding of RNAs and resolve RNAs that have misfolded, thereby ensuring that RNA is accessible for its biological functions.
Department of Biochemistry, Hughes Medical Institute, Stanford University, CA 94305-5307.
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Herschlag
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Khosla
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Tsuchihashi
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Karpel
D000595:Amino Acid Sequence; D001483:Base Sequence; D002213:Capsid / Q000378:metabolism; D036022:Capsid Proteins; D002384:Catalysis; D015683:Gene Products, gag / Q000378:metabolism; D000076245:Heterogeneous Nuclear Ribonucleoprotein A1; D034481:Heterogeneous-Nuclear Ribonucleoprotein Group A-B; D034441:Heterogeneous-Nuclear Ribonucleoproteins; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D009842:Oligopeptides / Q000494:pharmacology; D016337:RNA, Catalytic / Q000378:metabolism* / Q000494:pharmacology; D016601:RNA-Binding Proteins / Q000378:metabolism* / Q000494:pharmacology; D012261:Ribonucleoproteins / Q000378:metabolism; D014764:Viral Proteins; D054301:gag Gene Products, Human Immunodeficiency Virus
D036022:Capsid Proteins; D015683:Gene Products, gag; D000076245:Heterogeneous Nuclear Ribonucleoprotein A1; D034481:Heterogeneous-Nuclear Ribonucleoprotein Group A-B; D034441:Heterogeneous-Nuclear Ribonucleoproteins; C071884:NCP7 protein, Human immunodeficiency virus 1; D009842:Oligopeptides; D016337:RNA, Catalytic; D016601:RNA-Binding Proteins; D012261:Ribonucleoproteins; D014764:Viral Proteins; D054301:gag Gene Products, Human Immunodeficiency Virus