Cleavage of Model Substrates by Arabidopsis thaliana PRORP1 Reveals New Insights into Its Substrate Requirements

Cleavage of Model Substrates by Arabidopsis thaliana PRORP1 Reveals New Insights into Its Substrate Requirements

Two broad classes of RNase P trim the 5' leader of precursor tRNAs (pre-tRNAs): ribonucleoprotein (RNP)- and proteinaceous (PRORP)-variants. These two RNase P types, which use different scaffolds for catalysis, reflect independent evolutionary paths. While the catalytic RNA-based RNP form is pr...

Full description

Bibliographic Details
Main Authors: Mao, Guanzhong, Chen, Tien-Hao, Srivastava, Abhishek S., Kosek, David, Biswas, Pradip K., Gopalan, Venkat, Kirsebom, Leif A.
Format: Online
Language:English
Published: Public Library of Science 2016
Online Access:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4975455/
id pubmed-4975455
recordtype oai_dc
spelling pubmed-49754552016-08-25 Cleavage of Model Substrates by Arabidopsis thaliana PRORP1 Reveals New Insights into Its Substrate Requirements Mao, Guanzhong Chen, Tien-Hao Srivastava, Abhishek S. Kosek, David Biswas, Pradip K. Gopalan, Venkat Kirsebom, Leif A. Research Article Two broad classes of RNase P trim the 5' leader of precursor tRNAs (pre-tRNAs): ribonucleoprotein (RNP)- and proteinaceous (PRORP)-variants. These two RNase P types, which use different scaffolds for catalysis, reflect independent evolutionary paths. While the catalytic RNA-based RNP form is present in all three domains of life, the PRORP family is restricted to eukaryotes. To obtain insights on substrate recognition by PRORPs, we examined the 5' processing ability of recombinant Arabidopsis thaliana PRORP1 (AtPRORP1) using a panel of pre-tRNASer variants and model hairpin-loop derivatives (pATSer type) that consist of the acceptor-T-stem stack and the T-/D-loop. Our data indicate the importance of the identity of N-1 (the residue immediately 5' to the cleavage site) and the N-1:N+73 base pair for cleavage rate and site selection of pre-tRNASer and pATSer. The nucleobase preferences that we observed mirror the frequency of occurrence in the complete suite of organellar pre-tRNAs in eight algae/plants that we analyzed. The importance of the T-/D-loop in pre-tRNASer for tight binding to AtPRORP1 is indicated by the 200-fold weaker binding of pATSer compared to pre-tRNASer, while the essentiality of the T-loop for cleavage is reflected by the near-complete loss of activity when a GAAA-tetraloop replaced the T-loop in pATSer. Substituting the 2'-OH at N-1 with 2'-H also resulted in no detectable cleavage, hinting at the possible role of this 2'-OH in coordinating Mg2+ ions critical for catalysis. Collectively, our results indicate similarities but also key differences in substrate recognition by the bacterial RNase P RNP and AtPRORP1: while both forms exploit the acceptor-T-stem stack and the elbow region in the pre-tRNA, the RNP form appears to require more recognition determinants for cleavage-site selection. Public Library of Science 2016-08-05 /pmc/articles/PMC4975455/ /pubmed/27494328 http://dx.doi.org/10.1371/journal.pone.0160246 Text en © 2016 Mao et al http://creativecommons.org/licenses/by/4.0/ This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
repository_type Open Access Journal
institution_category Foreign Institution
institution US National Center for Biotechnology Information
building NCBI PubMed
collection Online Access
language English
format Online
author Mao, Guanzhong
Chen, Tien-Hao
Srivastava, Abhishek S.
Kosek, David
Biswas, Pradip K.
Gopalan, Venkat
Kirsebom, Leif A.
spellingShingle Mao, Guanzhong
Chen, Tien-Hao
Srivastava, Abhishek S.
Kosek, David
Biswas, Pradip K.
Gopalan, Venkat
Kirsebom, Leif A.
Cleavage of Model Substrates by Arabidopsis thaliana PRORP1 Reveals New Insights into Its Substrate Requirements
author_facet Mao, Guanzhong
Chen, Tien-Hao
Srivastava, Abhishek S.
Kosek, David
Biswas, Pradip K.
Gopalan, Venkat
Kirsebom, Leif A.
author_sort Mao, Guanzhong
title Cleavage of Model Substrates by Arabidopsis thaliana PRORP1 Reveals New Insights into Its Substrate Requirements
title_short Cleavage of Model Substrates by Arabidopsis thaliana PRORP1 Reveals New Insights into Its Substrate Requirements
title_full Cleavage of Model Substrates by Arabidopsis thaliana PRORP1 Reveals New Insights into Its Substrate Requirements
title_fullStr Cleavage of Model Substrates by Arabidopsis thaliana PRORP1 Reveals New Insights into Its Substrate Requirements
title_full_unstemmed Cleavage of Model Substrates by Arabidopsis thaliana PRORP1 Reveals New Insights into Its Substrate Requirements
title_sort cleavage of model substrates by arabidopsis thaliana prorp1 reveals new insights into its substrate requirements
description Two broad classes of RNase P trim the 5' leader of precursor tRNAs (pre-tRNAs): ribonucleoprotein (RNP)- and proteinaceous (PRORP)-variants. These two RNase P types, which use different scaffolds for catalysis, reflect independent evolutionary paths. While the catalytic RNA-based RNP form is present in all three domains of life, the PRORP family is restricted to eukaryotes. To obtain insights on substrate recognition by PRORPs, we examined the 5' processing ability of recombinant Arabidopsis thaliana PRORP1 (AtPRORP1) using a panel of pre-tRNASer variants and model hairpin-loop derivatives (pATSer type) that consist of the acceptor-T-stem stack and the T-/D-loop. Our data indicate the importance of the identity of N-1 (the residue immediately 5' to the cleavage site) and the N-1:N+73 base pair for cleavage rate and site selection of pre-tRNASer and pATSer. The nucleobase preferences that we observed mirror the frequency of occurrence in the complete suite of organellar pre-tRNAs in eight algae/plants that we analyzed. The importance of the T-/D-loop in pre-tRNASer for tight binding to AtPRORP1 is indicated by the 200-fold weaker binding of pATSer compared to pre-tRNASer, while the essentiality of the T-loop for cleavage is reflected by the near-complete loss of activity when a GAAA-tetraloop replaced the T-loop in pATSer. Substituting the 2'-OH at N-1 with 2'-H also resulted in no detectable cleavage, hinting at the possible role of this 2'-OH in coordinating Mg2+ ions critical for catalysis. Collectively, our results indicate similarities but also key differences in substrate recognition by the bacterial RNase P RNP and AtPRORP1: while both forms exploit the acceptor-T-stem stack and the elbow region in the pre-tRNA, the RNP form appears to require more recognition determinants for cleavage-site selection.
publisher Public Library of Science
publishDate 2016
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4975455/
_version_ 1613622564198809600

Similar Items