Morpholino oligonucleotides do not participate perfectly in standard Watson-Crick complexes with RNA

RNA. 2010 Nov;16(11):2218-25. doi: 10.1261/rna.2256610. Epub 2010 Sep 3.

Abstract

RNase P from E. coli will cleave a RNA at a site designated in a complex with an external guide sequence (EGS). The location of the site is determined by the Watson-Crick complementary sequence that can be formed between the RNA and the EGS. Morpholino oligonucleotides (PMOs) that have the same base sequences as any particular EGS will not direct cleavage by RNase P of the target RNA at the expected site in three mRNAs. Instead, cleavage occurs at a secondary site that does not correspond exactly to the expected Watson-Crick sequence in the PMO. This cleavage in the mRNA for a drug resistance gene, CAT mRNA, is at least second order in the concentration of the PMOs, but the mechanism is not understood yet and might be more complicated than a simple second-order reaction. EGSs and PMOs inhibit the reactions of each other effectively in a competitive fashion. A basic peptide attached to the PMO (PPMO) is more effective because of its binding properties to the mRNA as a substrate. However, a PMO is just as efficient as a PPMO on a mRNA that is mutated so that the canonical W-C site has been altered. The altered mRNA is not recognizable by effective extensive W-C pairing to an EGS or PMO. The complex of a PMO on a mutated mRNA as a substrate shows that the dimensions of the modified oligonucleotide cannot be the same as a naked piece of single-stranded RNA.

Publication types

  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Base Pairing*
  • Kinetics
  • Mutation
  • Oligonucleotides / chemistry*
  • Oligonucleotides / metabolism
  • RNA, Messenger / chemistry*
  • RNA, Messenger / genetics
  • RNA, Messenger / metabolism
  • Substrate Specificity

Substances

  • Oligonucleotides
  • RNA, Messenger