Multiple binding modes of substrate to the catalytic RNA subunit of RNase P from Escherichia coli

Krummel, Daniel A. Pomeranz; Altman, Sidney

doi:10.1017/s1355838299990416

Cited by 30 publications

(11 citation statements)

References 43 publications

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“…The processing of precursor tRNA molecules by M1 RNA is signi®cantly enhanced by C5 protein in vitro (Altman & Kirsebom, 1999), consistent with the observation that RNase P functions as an RNP complex in vivo (Sakano et al, 1974). Recent studies indicate that the protein subunit enhances the af®nity of the RNase P holoenzyme for the substrate perhaps by promoting speci®c interactions with the leader sequence in the substrate (Crary et al, 1998;Kurz et al, 1998;Niranjanakumari et al, 1998;Krummel & Altman, 1999).…”

Section: Introductionsupporting

confidence: 59%

Mapping RNA-protein interactions in ribonuclease P from Escherichia coli using disulfide-linked EDTA-fe 1 1Edited by K. Nagai

Biswas

Ledman

Fox

et al. 2000

Journal of Molecular Biology

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Section: Introductionsupporting

confidence: 59%

Mapping RNA-protein interactions in ribonuclease P from Escherichia coli using disulfide-linked EDTA-fe 1 1Edited by K. Nagai

Biswas

Ledman

Fox

et al. 2000

Journal of Molecular Biology

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“…Our archaeal RNase P data could be interpreted by a framework similar to that used to describe the role of the bacterial RPP ( 60 ; Scheme I and Supplementary Figure S7 ). Various kinetic and structural studies on bacterial RNase P have indicated that subsequent to substrate binding, a conformational change converts ES to ES* (defined by the equilibrium constant K conf ) and optimally positions the pre-tRNA and catalytic Mg 2+ ions for cleavage (at rate k c ) ( 58–61 ). Adding to a growing body of evidence supporting such a two-step mechanism, data from recent stopped-flow kinetic studies ( 61 ) confirm an initial bi-molecular collision (E + S → ES) followed by a uni-molecular conformational change (ES→ES*).…”

Section: Discussionmentioning

confidence: 99%

Dissecting functional cooperation among protein subunits in archaeal RNase P, a catalytic ribonucleoprotein complex

Chen¹,

Pulukkunat²,

Cho³

et al. 2010

Nucleic Acids Research

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RNase P catalyzes the Mg2+-dependent 5′-maturation of precursor tRNAs. Biochemical studies on the bacterial holoenzyme, composed of one catalytic RNase P RNA (RPR) and one RNase P protein (RPP), have helped understand the pleiotropic roles (including substrate/Mg2+ binding) by which a protein could facilitate RNA catalysis. As a model for uncovering the functional coordination among multiple proteins that aid an RNA catalyst, we use archaeal RNase P, which comprises one catalytic RPR and at least four RPPs. Exploiting our previous finding that these archaeal RPPs function as two binary RPP complexes (POP5•RPP30 and RPP21•RPP29), we prepared recombinant RPP pairs from three archaea and established interchangeability of subunits through homologous/heterologous assemblies. Our finding that archaeal POP5•RPP30 reconstituted with bacterial and organellar RPRs suggests functional overlap of this binary complex with the bacterial RPP and highlights their shared recognition of a phylogenetically-conserved RPR catalytic core, whose minimal attributes we further defined through deletion mutagenesis. Moreover, single-turnover kinetic studies revealed that while POP5•RPP30 is solely responsible for enhancing the RPR’s rate of precursor tRNA cleavage (by 60-fold), RPP21•RPP29 contributes to increased substrate affinity (by 16-fold). Collectively, these studies provide new perspectives on the functioning and evolution of an ancient, catalytic ribonucleoprotein.

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“…The scheme below was used to explain the role of the bacterial RPP in catalysis. Subsequent to substrate binding, a conformational change from ES to ES*, which helps position the ptRNA and catalytic metal ions optimally for cleavage, had already been proposed based on results from various kinetic and structural studies ( 38 , 39 ). Because the bacterial RPR's k obs with tight-binding, consensus ptRNAs increases only three fold by the RPP, the possibility of the RPP contributing functional groups to catalysis and thereby increasing k c (the rate of the chemical step) was discounted.…”

Section: Discussionmentioning

confidence: 99%

Studies on Methanocaldococcus jannaschii RNase P reveal insights into the roles of RNA and protein cofactors in RNase P catalysis

Pulukkunat

Gopalan

2008

Nucleic Acids Research

113

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Ribonuclease P (RNase P), a ribonucleoprotein (RNP) complex required for tRNA maturation, comprises one essential RNA (RPR) and protein subunits (RPPs) numbering one in bacteria, and at least four in archaea and nine in eukarya. While the bacterial RPR is catalytically active in vitro, only select euryarchaeal and eukaryal RPRs are weakly active despite secondary structure similarity and conservation of nucleotide identity in their putative catalytic core. Such a decreased archaeal/eukaryal RPR function might imply that their cognate RPPs provide the functional groups that make up the active site. However, substrate-binding defects might mask the ability of some of these RPRs, such as that from the archaeon Methanocaldococcus jannaschii (Mja), to catalyze precursor tRNA (ptRNA) processing. To test this hypothesis, we constructed a ptRNA-Mja RPR conjugate and found that indeed it self-cleaves efficiently (kobs, 0.15 min−1 at pH 5.5 and 55°C). Moreover, one pair of Mja RPPs (POP5-RPP30) enhanced kobs for the RPR-catalyzed self-processing by ∼100-fold while the other pair (RPP21-RPP29) had no effect; both binary RPP complexes significantly reduced the monovalent and divalent ionic requirement. Our results suggest a common RNA-mediated catalytic mechanism in all RNase P and help uncover parallels in RNase P catalysis hidden by plurality in its subunit make-up.

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Multiple binding modes of substrate to the catalytic RNA subunit of RNase P from Escherichia coli

Cited by 30 publications

References 43 publications

Mapping RNA-protein interactions in ribonuclease P from Escherichia coli using disulfide-linked EDTA-fe 1 1Edited by K. Nagai

Mapping RNA-protein interactions in ribonuclease P from Escherichia coli using disulfide-linked EDTA-fe 1 1Edited by K. Nagai

Dissecting functional cooperation among protein subunits in archaeal RNase P, a catalytic ribonucleoprotein complex

Studies on Methanocaldococcus jannaschii RNase P reveal insights into the roles of RNA and protein cofactors in RNase P catalysis

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