Cleavage of Model Substrates by Arabidopsis thaliana PRORP1 Reveals New Insights into Its Substrate Requirements
| Autor: | David Kosek, Guanzhong Mao, Tien-Hao Chen, Pradip K. Biswas, Leif A. Kirsebom, Abhishek Srivastava, Venkat Gopalan |
|---|---|
| Rok vydání: | 2016 |
| Předmět: |
0301 basic medicine
Chloroplasts Hydrolases lcsh:Medicine Plant Science Biochemistry Substrate Specificity Nucleobase RNA Precursors Ribozymes Magnesium RNA Small Interfering lcsh:Science Energy-Producing Organelles Ribonucleoprotein Multidisciplinary Ribozyme Läkemedelskemi Plants Enzymes Mitochondria Nucleic acids Chemistry Ribonucleoproteins RNA Plant Physical Sciences Transfer RNA Cellular Structures and Organelles Cellular Types Research Article Nucleases Base pair RNase P Stereochemistry Plant Cell Biology Arabidopsis Thaliana Fluorescence Polarization Brassica Bioenergetics Biology Research and Analysis Methods Cleavage (embryo) Catalysis Ribonuclease P 03 medical and health sciences Ribonucleases Model Organisms Plant and Algal Models Plant Cells DNA-binding proteins Botany Escherichia coli Non-coding RNA RNA Transfer Ser Biology and life sciences Arabidopsis Proteins lcsh:R Organisms Proteins Cell Biology RNase MRP 030104 developmental biology Enzymology biology.protein RNA Nucleic Acid Conformation lcsh:Q Medicinal Chemistry |
| Zdroj: | PLoS ONE PLoS ONE, Vol 11, Iss 8, p e0160246 (2016) |
| ISSN: | 1932-6203 |
| DOI: | 10.1371/journal.pone.0160246 |
| Popis: | 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-tRNA(Ser) 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-tRNA(Ser) 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-tRNA(Ser) for tight binding to AtPRORP1 is indicated by the 200-fold weaker binding of pATSer compared to pre-tRNA(Ser), 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. De två första författarna delar förstaförfattarskapet. |
| Databáze: | OpenAIRE |
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