Structure-based cleavage mechanism of Thermus thermophilus Argonaute DNA guide strand-mediated DNA target cleavage

G. Sheng, H. Zhao, J. Wang, Y. Rao, W. Tian, D.C. Swarts, J. van der Oost, D.J. Patel, Y. Wang

Research output: Contribution to journalArticleAcademicpeer-review

83 Citations (Scopus)

Abstract

We report on crystal structures of ternary Thermus thermophilus Argonaute (TtAgo) complexes with 5'-phosphorylated guide DNA and a series of DNA targets. These ternary complex structures of cleavage-incompatible, cleavage-compatible, and postcleavage states solved at improved resolution up to 2.2 Å have provided molecular insights into the orchestrated positioning of catalytic residues, a pair of Mg2+ cations, and the putative water nucleophile positioned for in-line attack on the cleavable phosphate for TtAgo-mediated target cleavage by a RNase H-type mechanism. In addition, these ternary complex structures have provided insights into protein and DNA conformational changes that facilitate transition between cleavage-incompatible and cleavage-compatible states, including the role of a Glu finger in generating a cleavage-competent catalytic Asp-Glu-Asp-Asp tetrad. Following cleavage, the seed segment forms a stable duplex with the complementary segment of the target strand
Original languageEnglish
Pages (from-to)652-657
JournalProceedings of the National Academy of Sciences of the United States of America
Volume111
Issue number2
DOIs
Publication statusPublished - 2014

Keywords

  • crystal-structure
  • rna recognition
  • silencing complex
  • substrate-specificity
  • slicer activity
  • piwi protein
  • paz domain
  • human risc
  • endonuclease
  • interference

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