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Visualizing the RNase H1 catalyzed cleavage of RNA using time-resolved X-ray crystallography

Friday, November 08, 2013 — Poster Session IV

2:00 p.m. – 4:00 p.m.

FAES Academic Center (Upper-Level Terrace)




  • NL Samara
  • W Yang


RNase H1 is an endonuclease that specifically recognizes and cleaves RNA strands in RNA/DNA hybrids. The enzyme exists in various life forms and has many cellular functions, including the cleavage of RNA primers from Okazaki fragments during replication. Many other proteins contain an RNase H1 catalytic domain, including HIV reverse transcriptase, where RNase H1 is needed for the conversion of a single stranded RNA viral genome into double stranded DNA. While studies suggest that RNase H1 mediated catalysis occurs by a two-metal ion mechanism, the reaction itself has never been visualized. Using time resolved X-ray crystallography, the hydrolysis of RNA can be followed. RNase H1 was co-crystallized with a RNA/DNA hybrid substrate in the absence of Mg2+. The reaction was initiated by exposing crystals to Mg2+ and stopped by freezing at specific time points for structural analysis. Initial structures from reactions conducted at various pHs reveal novel and unexpected characteristics of RNase H1 mediated RNA hydrolysis.

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