title: Binding and cleavage of unstructured RNA by nuclear RNase P.
abstract: Ribonuclease P (RNase P) is an essential endoribonuclease for which the best-characterized function is processing the 5' leader of pre-tRNAs. Compared to bacterial RNase P, which contains a single small protein subunit and a large catalytic RNA subunit, eukaryotic nuclear RNase P is more complex, containing nine proteins and an RNA subunit in Saccharomyces cerevisiae. Consistent with this, nuclear RNase P has been shown to possess unique RNA binding capabilities. To understand the unique molecular recognition of nuclear RNase P, the interaction of S. cerevisiae RNase P with single-stranded RNA was characterized. Unstructured, single-stranded RNA inhibits RNase P in a size-dependent manner, suggesting that multiple interactions are required for high affinity binding. Mixed-sequence RNAs from protein-coding regions also bind strongly to the RNase P holoenzyme. However, in contrast to poly(U) homopolymer RNA that is not cleaved, a variety of mixed-sequence RNAs have multiple preferential cleavage sites that do not correspond to identifiable consensus structures or sequences. In addition, pre-tRNA(Tyr), poly(U)(50) RNA, and mixed-sequence RNA cross-link with purified RNase P in the RNA subunit Rpr1 near the active site in "Conserved Region I," although the exact positions vary. Additional contacts between poly(U)(50) and the RNase P proteins Rpr2p and Pop4p were identified. We conclude that unstructured RNAs interact with multiple protein and RNA contacts near the RNase P RNA active site, but that cleavage depends on the nature of interaction with the active site.
authors.affiliation: Department of Biological Chemistry, University of Michigan, Ann Arbor, Michigan 48109-0606, USA.
authors.forename: Michael C
authors.initials: MC
authors.lastname: Marvin
authors.forename: Scott C
authors.initials: SC
authors.lastname: Walker
authors.forename: Carol A
authors.initials: CA
authors.lastname: Fierke
authors.forename: David R
authors.initials: DR
authors.lastname: Engelke
mesh_terms: D001483:Base Sequence; D002467:Cell Nucleus / Q000201:enzymology; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D011485:Protein Binding; D022661:RNA, Untranslated / Q000737:chemistry / Q000378:metabolism*; D043262:Ribonuclease P / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000201:enzymology*
chemical_list: D022661:RNA, Untranslated; D043262:Ribonuclease P