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Query: EC:3.1.26.5 (RNase P)
 1,348 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Two Bacillus subtilis tRNA(His) precursors (Green, C. J., and Vold, B. S. (1988) J. Biol. Chem. 263, 652-657) were processed by Escherichia coli RNase P in the presence of varying [Mg2+]. The wild type precursor was processed under all conditions to afford a single tRNA product containing 8 base pairs in the acceptor stem. In contrast, the position of processing of a mutant tRNA(His) precursor (containing a G27----A27 alteration) was shown to be condition-dependent. Processing occurred at A27 under conditions consistent with formation of an A27-C100 base pair in the acceptor stem but at G28 under conditions that disfavored base pair formation. The ability to control the site of RNase P-mediated tRNA precursor processing is unprecedented and permits analysis of the chemical factors that promote processing.
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PMID:Control of the position of RNase P-mediated transfer RNA precursor processing. 211 Jan 59

Ribonuclease P is the endonuclease that removes the leader fragments from the 5'-ends of precursor tRNAs. The enzyme isolated from eubacteria contains a catalytic RNA subunit. RNAs also copurify with eukaryotic RNase P, although catalysis by those RNAs has not been demonstrated. This paper reports the isolation and characterization of ribonuclease P from the thermoacidophilic archaebacterium Sulfolobus solfataricus. Archaebacteria are a primary evolutionary lineage, distinct from both eukaryotes and eubacteria. Ribonuclease P of S. solfataricus has reaction component requirements and a Km for substrate tRNA (2.5 X 10(-7) M) that are roughly similar to those reported for eubacterial and eukaryotic ribonuclease P. The temperature optimum for the reaction is 77 degrees C, reflecting the thermophilic character of the organism. The enzyme activity is not affected by treatment with micrococcal nuclease, suggesting that there is no RNA subunit or that it is protected from nuclease action. The density of the enzyme in cesium sulfate equilibrium density gradients is 1.27 g/ml, which is similar to that of protein. However, several RNAs between 200 and 400 nucleotides in size copurify with the enzyme activity on the density gradients, and one of them remains after micrococcal nuclease treatment. These properties of the S. solfataricus enzyme are compared with those of ribonuclease P from eukaryotes and eubacteria.
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PMID:Characterization of ribonuclease P from the archaebacterium Sulfolobus solfataricus. 211 85

Several modified nucleosides were introduced during in vitro RNA synthesis into a pre-tRNA(Ser). The pre-tRNAs were used as substrates for RNase P enzymes. No effects were observed with biotin-8-ATP or [alpha-S]-GPT, whereas with m7GTP, the cleavage reaction was completely inhibited. Analysis of pre-tRNAs which contained m7G at various positions has revealed a single base at the 5'-end of the acceptor stem where this modification absolutely prevents cleavage by catalytic M1 RNA, eukaryotic and prokaryotic RNase P holoenzymes. These results suggest that a critical contact must be made between pre-tRNA substrate and enzyme/ribozyme or that the approach of the potential cleaving agent (a positive magnesium ion) is made impossible by the positive charge at N-7 of the guanosine. In addition, we have shown that a pre-tRNA containing only m7G's can still form a complex with M1 RNA in a gel retardation assay.
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PMID:The methylation of one specific guanosine in a pre-tRNA prevents cleavage by RNase P and by the catalytic M1 RNA. 217 70

In a previous study it was shown that RNase P from E. coli cleaves the tRNA-like structure of turnip yellow mosaic virus (TYMV) RNA in vitro (Guerrier-Takada et al. (1988) Cell, 53, 267-272). Cleavage takes place at the 3' side of the loop that crosses the deep groove of the pseudoknot structure present in the aminoacyl acceptor domain. In the present study fragments of TYMV RNA with mutations in the pseudoknot, generated by transcription in vitro, were tested for susceptibility to cleavage by RNase P. Changes in the specificity with respect to the site of cleavage and decreases in the rate of cleavage were observed with most of these substrates. The behaviour of various mutants in the reaction catalyzed by RNase P is in agreement with the present model of the TYMV RNA pseudoknot (Dumas et al. (1987), J. Biomol. Struct. Dyn. 263, 652-657). Base substitutions in the loop that crosses the shallow groove of the pseudoknot structure resulted, however, in an unexpected decrease in the rate of cleavage, probably due to conformational changes in the substrates. Studies on other tRNA-like structures revealed an important role in the reaction with RNase P for both the nucleotide at the 3' side of the loop that spans the deep groove and the nucleotide at position 4, which correspond to positions--1 and 73, respectively, in tRNA precursors.
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PMID:Interaction of RNase P from Escherichia coli with pseudoknotted structures in viral RNAs. 219 61

Several "dimeric" tRNA molecules were constructed as potential substrates for ribonuclease P (RNase P) and for M1 RNA, the catalytic subunit of RNase P. Construction was affected by the T4 RNA ligase-mediated coupling of a mature Escherichia coli tRNA (acceptor substrate) and nucleotides 1-36 of yeast tRNAPhe (donor substrate), followed by annealing of the 3'-half of yeast tRNAPhe (nucleotides 38-76). E. coli RNase P and M1 RNA were both found to cleave the dimeric tRNA precursor model constructed from E. coli tRNAPhe (5'-tRNA) and yeast tRNAPhe (3'-tRNA) in a reaction that was dependent on the presence of the annealed 3'-half molecule derived from yeast tRNAPhe, or on some conformation imposed by the presence of this species; the product had the same mobility as authentic E. coli tRNAPhe on a polyacrylamide gel. By utilizing tRNA precursor models radiolabeled at phosphodiesters immediately preceding or following the putative site of processing, cleavage of the substrate by both M1 RNA and the holoenzyme was demonstrated to occur at the expected phosphate ester linkage. The results obtained here suggest that the endonucleolytic separation of two tRNAs by RNase P is dependent on one or more structural features in the 3'-half of the 3'-tRNA, and thus are consistent with the report of McClain et al. (McClain, W. H., Guerrier-Takada, C., and Altman, S. (1987) Science 238, 527-530) that identifies the T stem and loop as a possible recognition site.
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PMID:Construction and processing of transfer RNA precursor models. 226 40

A synthetic tRNA precursor analog containing the structural elements of Escherichia coli tRNA(Phe) was characterized as a substrate for E. coli ribonuclease P and for M1 RNA, the catalytic RNA subunit. Processing of the synthetic precursor exhibited a Mg2+ dependence quite similar to that of natural tRNA precursors such as E. coli tRNA(Tyr) precursor. It was found that Sr2+, Ca2+, and Ba2+ ions promoted processing of the dimeric precursor at Mg2+ concentrations otherwise insufficient to support processing; very similar behavior was noted for E. coli tRNA(Tyr). As noted previously for natural tRNA precursors, the absence of the 3'-terminal CA sequence in the synthetic precursor diminished the facility of processing of this substrate by RNase P and M1 RNA. A study of the Mg2+ dependence of processing of the synthetic tRNA dimeric substrate radiolabeled between C75 and A76 provided unequivocal evidence for an alteration in the actual site of processing by E. coli RNase P as a function of Mg2+ concentration. This property was subsequently demonstrated to obtain (Carter, B. J., Vold, B.S., and Hecht, S. M. (1990) J. Biol. Chem. 265, 7100-7103) for a mutant Bacillus subtilis tRNAHis precursor containing a potential A-C base pair at the end of the acceptor stem.
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PMID:Metal ion and substrate structure dependence of the processing of tRNA precursors by RNase P and M1 RNA. 226 41

Modified bases were introduced into pre-tRNAs during in vitro RNA synthesis or by chemical modification. These RNAs were used as substrates for the catalytic M1 RNA and the RNase P holoenzyme from Schizosaccharomyces pombe. The synthetic approach permitted the insertion of 100% m7GTP into pre-tRNAs and this resulted in complete inhibition of the specific 5' processing reactions. Partially modified RNAs were obtained by chemical modifications of purines and uridines in the pre-tRNAs. This allowed detailed analyses of specific bases excluded in the products. With pre-tRNA(Ser) and initiator pre-tRNA(Met), strong effects were observed in the T arm and weaker effects in the anticodon stem. Only minor base exclusions were detected in the acceptor stem of pre-tRNA(Ser) and in the D arm of pre-tRNA(Met).
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PMID:Substrate recognition by RNase P and by the catalytic M1 RNA: identification of possible contact points in pre-tRNAs. 234 11

A bank of temperature sensitive (ts-) mutants of Schizosaccharomyces pombe was screened for snRNA expression mutants using an oligodeoxynucleotide that recognizes U2 RNA. One mutant with a novel phenotype was identified that has reduced steady-state levels of the spliceosomal snRNAs U1, U2, U4, U5 and U6. In addition, the mutant exhibits a temperature-dependent accumulation of aberrant U2 and U4 transcripts elongated at their 3' end. The steady-state concentration of the RNA component of RNase P is also reduced in the mutant, whereas the amount of U3 RNA, 7SL RNA, tRNA, rRNA and mRNA are the same as wild-type. Pre-mRNA, pre-tRNA and U6 RNA precursor processing are impaired in the mutant. Genetic analysis demonstrates that the snRNA defects are tightly linked to the ts- growth defect and are recessive. We have named this mutant snm1 to indicate a defect in snRNA maintenance. The data on snm1 suggest that a single trans-acting factor is essential for the maintenance of steady-state levels of several snRNAs and for proper 3' end formation of U2 and U4 RNAs.
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PMID:A mutation in a single gene of Schizosaccharomyces pombe affects the expression of several snRNAs and causes defects in RNA processing. 240 30

M1 RNA, the RNA subunit of ribonuclease P from Escherichia coli, can under certain conditions catalytically cleave precursors to tRNA in the absence of C5, the protein moiety of RNase P. M1 RNA itself is not cleaved during the reaction, nor does it form any covalent bonds with its substrate. Only magnesium and, to a lesser extent, manganese ions can function at the catalytic center of M1 RNA. Several other ions either inhibit the binding of magnesium ion at the active site or function as structural counterions. The reaction rate of cleavage of precursors to tRNAs by M1 RNA is enhanced in the presence of poly-(ethylene glycol) or 2-methyl-2,4-pentanediol. Many aspects of the reaction catalyzed by M1 RNA are compatible with a mechanism in which phosphodiester bond cleavage is mediated by metal ion.
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PMID:Metal ion requirements and other aspects of the reaction catalyzed by M1 RNA, the RNA subunit of ribonuclease P from Escherichia coli. 242 12

Until the discovery of catalytic RNAs, first the self-splicing intron in Tetrahymena and then the bacterial RNAse P, cellular enzymes had always seemed to be protein in nature. The recognition that RNA can catalytically make and break phosphodiester bonds simplifies some of the assumptions required of a rudimentary self-replicating entity. Available information on the chemistry of RNA-catalyzed reactions is reviewed, with particular attention to self-splicing introns and tRNA processing by RNase P. An explicit model for a self-replicating RNA is described. The model postulates a nucleotide binding/polymerization site in the RNA, and takes advantage of intrinsic fluidity in RNA higher order structure to dissociate parent and progeny complementary strands.
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PMID:RNA catalysis and the origin of life. 242 41


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