Strict conformational demands of RNA cleavage in bulgeloops created by peptidyl-oligonucleotide conjugates

Research output: Contribution to journalArticlepeer-review

  • External authors:
  • Yaroslav Staroseletz
  • Bahareh Amirloo
  • Aled Williams
  • Alexander Lomzov
  • David Clarke
  • Tom Brown
  • Marina A Zenkova

Abstract

Potent knockdown of pathogenic RNA in vivo is an urgent health need unmet by both small-molecule and biologic drugs. ‘Smart’ supramolecular assembly of catalysts offers precise recognition and potent destruction of targeted RNA, hitherto not found in nature. Peptidyl-oligonucleotide ribonucleases are here chemically-engineered to create and attack bulge-loop regions upon
hybridisation to target RNA. Catalytic peptide was incorporated either via a centrally-modified nucleotide (Type 1) or through an abasic sugar residue (Type 2) within the RNA-recognition motif to reveal striking differences in biological performance and strict structural demands of ribonuclease activity. None of the Type 1 conjugates were catalytically active, whereas all Type 2 conjugates
cleaved RNA target in a sequence-specific manner, with up to 90% cleavage from 5 nucleotide bulgeloops (BC5-α and BC5L-β anomers) through multiple cuts, including in folds nearby. Molecular dynamics simulations provided structural explanation of accessibility of the RNA cleavage sites to the peptide with adoption of an “in-line” attack conformation for catalysis. Hybridisation assays and enzymatic probing with RNases illuminated how RNA binding specificity and dissociation after cleavage can be balanced to permit turnover of the catalytic reaction. This is an essential requirement for inactivation of multiple copies of disease-associated RNA and therapeutic efficacy.

Bibliographical metadata

Original languageEnglish
JournalNucleic Acids Res
Publication statusAccepted/In press - 7 Sep 2020