Human ribonuclease P: Subunits, function, and intranuclear localization

Jarrous, Nayef

doi:10.1017/s1355838202011184

Cited by 93 publications

(75 citation statements)

References 82 publications

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“…Furthermore, RNase P-catalyzed cleavage has also been suggested to play a role in mRNA degradation and regulation of gene expression in vivo . In most organisms, RNase P consists of a catalytically active RNA subunit (PRNA) associated with one or more protein cofactors (P proteins) essential for in vivo activity (Hall and Brown 2002;Jarrous 2002;Hsieh et al 2004;Evans et al 2006;Walker and Engelke 2006;Gopalan 2007;Jarrous and Reiner 2007). Given the essential and universal role of this enzyme, it is fundamentally important to understand the mechanisms of molecular recognition and catalysis used by RNase P.…”

Section: Introductionmentioning

confidence: 99%

Conformational change in the Bacillus subtilis RNase P holoenzyme–pre-tRNA complex enhances substrate affinity and limits cleavage rate

Hsieh¹,

Fierke²

2009

RNA

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Ribonuclease P (RNase P) is a ribonucleoprotein complex that catalyzes the 59 maturation of precursor tRNAs. To investigate the mechanism of substrate recognition in this enzyme, we characterize the thermodynamics and kinetics of Bacillus subtilis pretRNA Asp binding to B. subtilis RNase P holoenzyme using fluorescence techniques. Time courses for fluorescein-labeled pre-tRNA binding to RNase P are biphasic in the presence of both Ca(II) and Mg(II), requiring a minimal two-step association mechanism. In the first step, the apparent bimolecular rate constant for pre-tRNA associating with RNase P has a value that is near the diffusion limit and is independent of the length of the pre-tRNA leader. Following formation of the initial enzyme-substrate complex, a unimolecular step enhances the overall affinity of pre-tRNA by eight-to 300-fold as the length of the leader sequence increases from 2 to 5 nucleotides. This increase in affinity is due to a decrease in the reverse rate constant for the conformational change that correlates with the formation of an optimal leader-protein interaction in the RNase P holoenzyme-pre-tRNA complex. Furthermore, the forward rate constant for the conformational change becomes rate limiting for cleavage under single-turnover conditions at high pH, explaining the origin of the observed apparent pK a in the RNase P-catalyzed cleavage reaction. These data suggest that a conformational change in the RNase P d pre-tRNA complex is coupled to the interactions between the 59 leader and P protein and aligns essential functional groups at the cleavage active site to enhance efficient cleavage of pre-tRNA.

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Section: Introductionmentioning

confidence: 99%

Conformational change in the Bacillus subtilis RNase P holoenzyme–pre-tRNA complex enhances substrate affinity and limits cleavage rate

Hsieh¹,

Fierke²

2009

RNA

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“…Several human proteins (corresponding to homologs of Pop1, Pop3, Pop4, and Pop8 in yeast) were shown to accumulate in the nucleolus, suggesting that they might be involved in localization of RNase MRP and P (Jarrous et al 1999;van Eenennaam et al 2001). Several overexpressed human RNase P proteins were shown to selectively bind to precursor tRNA, suggesting that they might be involved in interaction with the substrate in the holoenzyme (Jarrous 2002). Filter binding assays (Aspinall et al 2007) suggested that individually expressed yeast proteins Pop4, Pop6, Rmp1, and Snm1 might interact with the RNase MRP pre-rRNA substrate.…”

Section: Introductionmentioning

confidence: 99%

Footprinting analysis demonstrates extensive similarity between eukaryotic RNase P and RNase MRP holoenzymes

Esakova

Perederina²,

Quan

et al. 2008

RNA

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Eukaryotic ribonuclease (RNase) P and RNase MRP are evolutionary related RNA-based enzymes involved in metabolism of various RNA molecules, including tRNA and rRNA. In contrast to the closely related eubacterial RNase P, which is comprised of an RNA component and a single small protein, these enzymes contain multiple protein components. Here we report the results of footprinting studies performed on purified Saccharomyces cerevisiae RNase MRP and RNase P holoenzymes. The results identify regions of the RNA components affected by the protein moiety, suggest a role of the proteins in stabilization of the RNA fold, and point to substantial similarities between the two evolutionary related RNA-based enzymes.

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“…Eukaryotic RNase P has a very similar RNA moiety, but the protein complement is far more complex than that of the bacterial enzyme. Nine-and 10-protein subunits have been identified in the nuclear RNase P from yeast and humans, respectively (Chamberlain et al 1998;Jarrous 2002). The eukaryotic RNase P RNA appears to have lost the ability to cleave pre-tRNAs in the absence of the protein subunits.…”

Section: Introductionmentioning

confidence: 99%

Functional characterization of the conserved amino acids in Pop1p, the largest common protein subunit of yeast RNases P and MRP

Xiao

Hsieh

Nugent

et al. 2006

RNA

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RNase P and RNase MRP are ribonucleoprotein enzymes required for 59-end maturation of precursor tRNAs (pre-tRNAs) and processing of precursor ribosomal RNAs, respectively. In yeast, RNase P and MRP holoenzymes have eight protein subunits in common, with Pop1p being the largest at >100 kDa. Little is known about the functions of Pop1p, beyond the fact that it binds specifically to the RNase P RNA subunit, RPR1 RNA. In this study, we refined the previous Pop1 phylogenetic sequence alignment and found four conserved regions. Highly conserved amino acids in yeast Pop1p were mutagenized by randomization and conditionally defective mutations were obtained. Effects of the Pop1p mutations on pre-tRNA processing, pre-rRNA processing, and stability of the RNA subunits of RNase P and MRP were examined. In most cases, functional defects in RNase P and RNase MRP in vivo were consistent with assembly defects of the holoenzymes, although moderate kinetic defects in RNase P were also observed. Most mutations affected both pre-tRNA and pre-rRNA processing, but a few mutations preferentially interfered with only RNase P or only RNase MRP. In addition, one temperature-sensitive mutation had no effect on either tRNA or rRNA processing, consistent with an additional role for RNase P, RNase MRP, or Pop1p in some other form. This study shows that the Pop1p subunit plays multiple roles in the assembly and function of of RNases P and MRP, and that the functions can be differentiated through the mutations in conserved residues.

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Human ribonuclease P: Subunits, function, and intranuclear localization

Cited by 93 publications

References 82 publications

Conformational change in the Bacillus subtilis RNase P holoenzyme–pre-tRNA complex enhances substrate affinity and limits cleavage rate

Conformational change in the Bacillus subtilis RNase P holoenzyme–pre-tRNA complex enhances substrate affinity and limits cleavage rate

Footprinting analysis demonstrates extensive similarity between eukaryotic RNase P and RNase MRP holoenzymes

Functional characterization of the conserved amino acids in Pop1p, the largest common protein subunit of yeast RNases P and MRP

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