SiRNA Carrying an (E)-vinylphosphonate Moiety at the 5΄ End of the Guide Strand Augments Gene Silencing by Enhanced Binding to Human Argonaute-2

Overview

Journal Nucleic Acids Res

Publisher Oxford University Press

Specialty Biochemistry

Date 2016 Dec 2

PMID 27903888

Citations 36

Authors

Elad Elkayam

Rubina Parmar

Christopher R Brown

Jennifer L Willoughby

Christopher S Theile

Muthiah Manoharan

Leemor Joshua-Tor

Affiliations

Soon will be listed here.

Abstract

Efficient gene silencing by RNA interference (RNAi) in vivo requires the recognition and binding of the 5΄- phosphate of the guide strand of an siRNA by the Argonaute protein. However, for exogenous siRNAs it is limited by the rapid removal of the 5΄- phosphate of the guide strand by metabolic enzymes. Here, we have determined the crystal structure of human Argonaute-2 in complex with the metabolically stable 5΄-(E)-vinylphosphonate (5΄-E-VP) guide RNA at 2.5-Å resolution. The structure demonstrates how the 5΄ binding site in the Mid domain of human Argonaute-2 is able to adjust the key residues in the 5΄-nucleotide binding pocket to compensate for the change introduced by the modified nucleotide. This observation also explains improved binding affinity of the 5΄-E-VP -modified siRNA to human Argonaute-2 in-vitro, as well as the enhanced silencing in the context of the trivalent N-acetylgalactosamine (GalNAc)-conjugated siRNA in mice relative to the un-modified siRNA.

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