Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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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)

Ribonuclease P (RNase P) is a ribonucleoprotein enzyme that cleaves precursor tRNA transcripts to give mature 5' ends. RNase P in eubacteria has a large, catalytic RNA subunit and a small protein subunit that are required for precursor tRNA cleavage in vivo. Although the eukaryotic holoenzymes have similar, large RNA subunits, previous work in a number of systems has suggested that the eukaryotic enzymes require a greater protein content. We have purified the Saccharomyces cerevisiae nuclear RNase P to apparent homogeneity, allowing the first comprehensive analysis of an unexpectedly complex subunit composition. Peptide sequencing by ion trap mass spectrometry identifies nine proteins that copurify with the nuclear RNase P RNA subunit, totaling 20-fold more protein than in the bacterial enzyme. All of these proteins are encoded by genes essential for RNase P activity and for cell viability. Previous genetic studies suggested that four proteins might be subunits of both RNase P and RNase MRP, the related rRNA processing enzyme. We demonstrate that all four of these proteins, Pop1p, Pop3p, Pop4p, and Rpp1p, are integral subunits of RNase P. In addition, four of the five newly identified protein subunits, Pop5p, Pop6p, Pop7p, and Pop8p, also appear to be shared between RNase P and RNase MRP. Only one polypeptide, Rpr2p, is unique to the RNase P holoenzyme by genetic depletion and immunoprecipitation studies. The large increase in the number of protein subunits over eubacterial RNase P is consistent with an increase in functional complexity in eukaryotes. The degree of structural similarity between nuclear RNase P and RNase MRP suggests that some aspects of their functions in pre-tRNA and pre-rRNA processing pathways might overlap or be coordinated.
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PMID:Purification and characterization of the nuclear RNase P holoenzyme complex reveals extensive subunit overlap with RNase MRP. 962 Aug 54

Ribonuclease P is a ribonucleoprotein complex that catalyzes the essential 5' maturation of all precursor tRNA molecules. The protein component both alters the conformation of the RNA component and enhances the substrate affinity and specificity. To facilitate biochemical and biophysical studies, the protein component of Bacillus subtilis ribonuclease P (RNase P) was overproduced in Escherichia coli using the native amino acid sequence with the initial 20 codons optimized for expression in E.coli . A simple purification procedure using consecutive cation exchange chromatography steps in the presence and absence of urea was developed to purify large quantities of P protein without contaminating nucleic acids. The identity of the recombinant protein as a cofactor of RNase P was established by its ability to stimulate the activity of the RNA component in low ionic strength buffer in a 1:1 stoichiometry. Circular dichroism studies indicate that P protein is a combination of alpha-helix and beta-sheet secondary structures and is quite stable, with a T m of 67 degrees C. The described methods facilitated the large scale purification of homogeneous, RNA-free P protein required for high resolution crystallographic analyses and may be useful for the preparation of other RNA binding proteins.
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PMID:Expression, purification and characterization of the recombinant ribonuclease P protein component from Bacillus subtilis. 962 4

Ribonuclease P (RNase P) is a ribonucleoprotein complex that catalyzes the formation of the mature 5' end of tRNA. To investigate the role of the protein component in enhancing the affinity of Bacillus subtilis RNase P for substrate (Kurz, J. C., Niranjanakumari, S., Fierke, C. A. (1998) Biochemistry 37, 2393), the kinetics and thermodynamics of binding and cleavage were analyzed for pre-tRNAAsp substrates containing 5' leader sequences of varying lengths (1-33 nucleotides). These data demonstrate that the cleavage rate constant catalyzed by the holoenzyme is not dependent on the leader length; however, the association rate constant for substrate binding to holoenzyme increases as the length of the leader increases, and this is reflected in enhanced substrate affinity of up to 4 kcal/mol. In particular, the protein component of RNase P stabilizes interactions with nucleotides at -2 and -5 in the 5' leader sequence of the pre-tRNA substrate. A 1 nucleotide leader decreases substrate affinity >/=15-fold compared to tRNAAsp due to ground-state destabilization of the enzyme-substrate complex. This destabilization is overcome by increasing the length of the leader to 2 nucleotides due to P RNA-pre-tRNA contacts that are stabilized by the P protein. The affinity of RNase P holoenzyme (but not RNA alone) for pre-tRNAAsp is further enhanced with a substrate containing a 5 nucleotide leader. These data indicate that novel direct or indirect interactions occur between the 5' leader sequence of pre-tRNAAsp and the protein component of RNase P.
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PMID:The protein component of Bacillus subtilis ribonuclease P increases catalytic efficiency by enhancing interactions with the 5' leader sequence of pre-tRNAAsp. 964 23

Ribonuclease P cleaves 5'-precursor sequences from pre-tRNAs. All cellular RNase P holoenzymes contain homologous RNA elements; the eucaryal RNase P RNA, in contrast to the bacterial RNA, is catalytically inactive in the absence of the protein component(s). To understand the function of eucaryal RNase P RNA, knowledge of its structure is needed. Considerable effort has been devoted to comparative studies of the structure of this RNA from diverse organisms, including eucaryotes, primarily fungi, but also a limited set of vertebrates. The substantial differences in the sequences and structures of the vertebrate RNAs from those of other organisms have made it difficult to align the vertebrate sequences, thus limiting comparative studies. To expand our understanding of the structure of diverse RNase P RNAs, we have isolated by PCR and sequenced 13 partial RNase P RNA genes from 11 additional vertebrate taxa representing most extant major vertebrate lineages. Based on a recently proposed structure of the core elements of RNase P RNA, we aligned the sequences and propose a minimum consensus secondary structure for the vertebrate RNase P RNA.
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PMID:Comparative structure analysis of vertebrate ribonuclease P RNA. 964 15

Ribonuclease P (RNase P) is the endoribonuclease that generates the mature 5'-ends of tRNA by removal of the 5'-leader elements of precursor-tRNAs. This enzyme has been characterized from representatives of all three domains of life (Archaea, Bacteria, and Eucarya) (1) as well as from mitochondria and chloroplasts. The cellular and mitochondrial RNase Ps are ribonucleoproteins, whereas the most extensively studied chloroplast RNase P (from spinach) is composed solely of protein. Remarkably, the RNA subunit of bacterial RNase P is catalytically active in vitro in the absence of the protein subunit (2). Although RNA-only activity has not been demonstrated for the archael, eucaryal, or mitochondrial RNAs, comparative sequence analysis has established that these RNAs are homologous (of common ancestry) to bacterial RNA. RNase P holoenzymes vary greatly in organizational complexity across the phylogenetic domains, primarily because of differences in the RNase P protein subunits: Mitochondrial, archaeal, and eucaryal holoenzymes contain larger, and perhaps more numerous, protein subunits than do the bacterial holoenzymes. However, that the nonbacterial RNase P RNAs retain significant structural similarity to their catalytically active bacterial counterparts indicates that the RNA remains the catalytic center of the enzyme.
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PMID:Ribonuclease P: unity and diversity in a tRNA processing ribozyme. 975 86

Ribonuclease P is responsible for the 5'-maturation of tRNA precursors. Ribonuclease P is a ribonucleoprotein, and in bacteria (and some Archaea) the RNA subunit alone is catalytically active in vitro, i.e. it is a ribozyme. The Ribonuclease P Database is a compilation of ribonuclease P sequences, sequence alignments, secondary structures, three-dimensional models and accessory information, available via the World Wide Web at the following URL: http://www.mbio.ncsu.edu/RNaseP/home .html
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PMID:The Ribonuclease P Database. 984 14

Ribonuclease P (RNase P) is a catalytic ribonucleoprotein (RNP) essential for tRNA biosynthesis. In Escherichia coli, this RNP complex is composed of a catalytic RNA subunit, M1 RNA, and a protein cofactor, C5 protein. Using the sulfhydryl-specific reagent (1-oxyl-2,2,5, 5-tetramethyl-Delta3-pyrroline-3-methyl)methanethiosulfonate (MTSL), we have introduced a nitroxide spin label individually at six genetically engineered cysteine residues (i.e., positions 16, 21, 44, 54, 66, and 106) and the native cysteine residue (i.e., position 113) in C5 protein. The spin label covalently attached to any protein is sensitive to structural changes in its microenvironment. Therefore, we expected that if the spin label introduced at a particular position in C5 protein was present at the RNA-protein interface, the electron paramagnetic resonance (EPR) spectrum of the spin label would be altered upon binding of the spin-labeled C5 protein to M1 RNA. The EPR spectra observed with the various MTSL-modified mutant derivatives of C5 protein indicate that the spin label attached to the protein at positions 16, 44, 54, 66, and 113 is immobilized to varying degrees upon addition of M1 RNA but not in the presence of a catalytically inactive, deletion derivative of M1 RNA. In contrast, the spin label attached to position 21 displays an increased mobility upon binding to M1 RNA. The results from this EPR spectroscopy-based approach together with those from earlier studies identify residues in C5 protein which are proximal to M1 RNA in the RNase P holoenzyme complex.
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PMID:Mapping RNA-protein interactions in ribonuclease P from Escherichia coli using electron paramagnetic resonance spectroscopy. 1002 48

Ribonuclease P is the endonuclease required for generating the mature tRNA 5'-end. The ribonucleoprotein character of this enzyme has now been proven in most organisms and organelles. Exceptions, however, are still the chloroplasts, plant nuclei and animal mitochondria where no associated RNAs have been detected to date. In contrast to the known RNA subunits, which are fairly well-conserved in size and structure among diverse phylogenetic groups, the protein contribution to the holoenzyme is highly variable in size and number of the individual components. The structure of the bacterial protein component has recently been solved. In contrast, the spatial arrangement of the multiple subunits in eukaryotic enzymes is still enigmatic. Substrate requirements of the enzymes or their catalytic RNA subunits are equally diverse, ranging from simple single domain mimics to an almost intact three-dimensional structure of the pre-tRNA substrate. As an example for an intermediate in the enzyme evolution, ribonuclease P from the Cyanophora paradoxa cyanelle will be discussed in more detail. This enzyme is unique, as it combines cyanobacterial and eukaryotic features in its function, subunit composition and holoenzyme topology.
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PMID:Ribonuclease P: the diversity of a ubiquitous RNA processing enzyme. 1037 Oct 40

Ribonuclease P is the enzyme responsible for removing the 5'-leader segment of precursor transfer RNAs in all organisms. All eukaryotic nuclear RNase Ps are ribonucleoproteins in which multiple protein components and a single RNA species are required for activity in vitro as well as in vivo. It is not known, however, which subunits participate directly in phosphodiester-bond hydrolysis. The RNA subunit of nuclear RNase P is evolutionarily related to its catalytically active bacterial counterpart, prompting speculation that in eukaryotes the RNA may be the catalytic component. In the bacterial RNase P reaction, Mg(II) is required to coordinate the nonbridging phosphodiester oxygen(s) of the scissile bond. As a consequence, bacterial RNase P cannot cleave pre-tRNA in which the pro-Rp nonbridging oxygen of the scissile bond is replaced by sulfur. In contrast, the RNase P reaction in plant chloroplasts is catalyzed by a protein enzyme whose mechanism does not involve Mg(II) coordinated by the pro-Rp oxygen. To determine whether the mechanism of nuclear RNase P resembles more closely an RNA- or a protein-catalyzed reaction, we analyzed the ability of Saccharomyces cerevisiae nuclear RNase P to cleave pre-tRNA containing a sulfur substitution of the pro-Rp oxygen at the cleavage site. Sulfur substitution at this position prohibits correct cleavage of pre-tRNA. Cleavage by eukaryotic RNase P thus depends on the presence of a thio-sensitive ligand to the pro-Rp oxygen of the scissile bond, and is consistent with a common, RNA-based mechanism for the bacterial and eukaryal enzymes.
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PMID:Evidence for an RNA-based catalytic mechanism in eukaryotic nuclear ribonuclease P. 1078 46

Ribonuclease P (RNase P) is the ribonucleoprotein enzyme that cleaves 5'-leader sequences from precursor-tRNAs. Bacterial and eukaryal RNase P RNAs differ fundamentally in that the former, but not the latter, are capable of catalyzing pre-tRNA maturation in vitro in the absence of proteins. An explanation of these functional differences will be assisted by a detailed comparison of bacterial and eukaryal RNase P RNA structures. However, the structures of eukaryal RNase P RNAs remain poorly characterized, compared to their bacterial and archaeal homologs. Hence, we have taken a phylogenetic-comparative approach to refine the secondary structures of eukaryal RNase P RNAs. To this end, 20 new RNase P RNA sequences have been determined from species of ascomycetous fungi representative of the genera Arxiozyma, Clavispora, Kluyveromyces, Pichia, Saccharomyces, Saccharomycopsis, Torulaspora, Wickerhamia, and Zygosaccharomyces. Phylogenetic-comparative analysis of these and other sequences refines previous eukaryal RNase P RNA secondary structure models. Patterns of sequence conservation and length variation refine the minimum-consensus model of the core eukaryal RNA structure. In comparison to bacterial RNase P RNAs, the eukaryal homologs lack RNA structural elements thought to be critical for both substrate binding and catalysis. Nonetheless, the eukaryal RNA retains the main features of the catalytic core of the bacterial RNase P. This indicates that the eukaryal RNA remains intrinsically a ribozyme.
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PMID:Phylogenetic-comparative analysis of the eukaryal ribonuclease P RNA. 1114 87


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