Structures of RNA 3′-phosphate cyclase bound to ATP reveal the mechanism of nucleotidyl transfer and metal-assisted catalysis Journal Article


Authors: Chakravarty, A. K.; Smith, P.; Shuman, S.
Article Title: Structures of RNA 3′-phosphate cyclase bound to ATP reveal the mechanism of nucleotidyl transfer and metal-assisted catalysis
Abstract: RNA 3′-phosphate cyclase (RtcA) synthesizes RNA 2′,3′ cyclic phosphate ends via three steps: reaction with ATP to form a covalent RtcA-(histidinyl-Nε)-AMP intermediate; transfer of adenylate to an RNA 3′-phosphate to form RNA(3′)pp(5′)A; and attack of the vicinal O2′ on the 3′-phosphorus to form a 2′,3′ cyclic phosphate and release AMP. Here we report the crystal structures of RtcA·ATP, RtcA·ATP·Mn 2+, and RtcA·ATP·Co 2+ substrate complexes and an RtcA·AMP product complex. Together with the structures of RtcA apoenzyme and the covalent RtcA-AMP intermediate, they illuminate the mechanism of nucleotidyl transfer, especially the stereochemical transitions at the AMP phosphate, the critical role of the metal in orienting the PP i leaving group of ATP during step 1, and the protein conformational switches that accompany substrate binding and product release. The octahedral metal complex of RtcA·ATP·Mn 2+ includes nonbridging oxygens from each of the ATP phosphates, two waters, and Glu14 as the sole RtcA component. Whereas the RtcA adenylylation step is metal-catalyzed, the subsequent steps in the cyclization pathway are metal-independent.
Keywords: unclassified drug; protein conformation; oxygen; protein; protein binding; escherichia coli; crystal structure; models, molecular; catalysis; adenosine triphosphate; cobalt; protein structure; cyclization; crystallization; enzyme release; phosphate; stereochemistry; ligases; covalent catalysis; adenosine phosphate; adenylation; x-ray diffraction; manganese; rna 3' phosphate cyclase; rna modification; glu14 protein; oligonucleotide probes; rna nucleotidyltransferases
Journal Title: Proceedings of the National Academy of Sciences of the United States of America
Volume: 108
Issue: 52
ISSN: 0027-8424
Publisher: National Academy of Sciences  
Date Published: 2011-12-27
Start Page: 21034
End Page: 21039
Language: English
DOI: 10.1073/pnas.1115560108
PROVIDER: scopus
PMCID: PMC3248511
PUBMED: 22167800
DOI/URL:
Notes: --- - "Export Date: 1 March 2012" - "CODEN: PNASA" - "Source: Scopus"
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  1. Stewart H Shuman
    546 Shuman
  2. Paul M C Smith
    21 Smith