Novel mechanism of RNA repair by RtcB via sequential 2′,3′- cyclic phosphodiesterase and 3′-phosphate/5′-hydroxyl ligation reactions Journal Article
| Authors: | Tanaka, N.; Chakravarty, A. K.; Maughan, B.; Shuman, S. |
| Article Title: | Novel mechanism of RNA repair by RtcB via sequential 2′,3′- cyclic phosphodiesterase and 3′-phosphate/5′-hydroxyl ligation reactions |
| Abstract: | RtcB enzymes are a newly discovered family of RNA ligases, implicated in tRNA splicing and other RNA repair reactions, that seal broken RNAs with 2′,3′-cyclic phosphate and 5′-OH ends. Parsimony and energetics would suggest a one-step mechanism for RtcB sealing via attack by the O5′ nucleophile on the cyclic phosphate, with expulsion of the ribose O2′ and generation of a 3′,5′-phosphodiester at the splice junction. Yet we find that RtcB violates Occam's razor, insofar as (i) it is adept at ligating 3′-monophosphate and 5′-OH ends; (ii) it has an intrinsic 2′,3′-cyclic phosphodiesterase activity. The 2′,3′-cyclic phosphodiesterase and ligase reactions both require manganese and are abolished by mutation of the RtcB active site. Thus, RtcB executes a unique two-step pathway of strand joining whereby the 2′,3′-cyclic phosphodiester end is hydrolyzed to a 3′-monophosphate, which is then linked to the 5′-OH end to form the splice junction. The energy for the 3′-phosphate ligase activity is provided by GTP, which reacts with RtcB in the presence of manganese to form a covalent RtcB-guanylate adduct. This adduct is sensitive to acid and hydroxylamine but resistant to alkali, consistent with a phosphoramidate bond. |
| Keywords: | controlled study; unclassified drug; nonhuman; animal cell; protein protein interaction; enzyme activity; rna; mutagenesis, site-directed; enzyme structure; guanosine triphosphate; enzymes; enzyme mechanism; surgical equipment; rna splicing; escherichia coli proteins; esters; chemical bond; enzyme active site; ligase reaction; ligases; protein hydrolysis; ligase activity; rna repair; manganese; covalent bond; active site; cyclic phosphates; monophosphates; phosphodiesters; rna ligase; splice junctions; protein rtcb; phosphodiesterase; hydroxylamine; guanosine phosphate; ligation reactions; occam's razor; phosphodiesterase activity; phosphodiesterases; phosphoramidates; amino acyl-trna synthetases; 2',3' cyclic phosphodiesterase; 3' monophosphate ligase; 5 hydroxyl ligase; phosphoramidic acid; nucleotidases |
| Journal Title: | Journal of Biological Chemistry |
| Volume: | 286 |
| Issue: | 50 |
| ISSN: | 0021-9258 |
| Publisher: | American Society for Biochemistry and Molecular Biology |
| Date Published: | 2011-12-16 |
| Start Page: | 43134 |
| End Page: | 43143 |
| Language: | English |
| DOI: | 10.1074/jbc.M111.302133 |
| PROVIDER: | scopus |
| PMCID: | PMC3234866 |
| PUBMED: | 22045815 |
| DOI/URL: | |
| Notes: | --- - "Export Date: 3 January 2012" - "CODEN: JBCHA" - "Source: Scopus" |
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