SNP Discrimination by Tolane-Modified Peptide Nucleic Acids: Application for the Detection of Drug Resistance in Pathogens

Molecules. 2020 Feb 11;25(4):769. doi: 10.3390/molecules25040769.

Abstract

During the treatment of viral or bacterial infections, it is important to evaluate any resistance to the therapeutic agents used. An amino acid substitution arising from a single base mutation in a particular gene often causes drug resistance in pathogens. Therefore, molecular tools that discriminate a single base mismatch in the target sequence are required for achieving therapeutic success. Here, we synthesized peptide nucleic acids (PNAs) derivatized with tolane via an amide linkage at the N-terminus and succeeded in improving the sequence specificity, even with a mismatched base pair located near the terminal region of the duplex. We assessed the sequence specificities of the tolane-PNAs for single-strand DNA and RNA by UV-melting temperature analysis, thermodynamic analysis, an in silico conformational search, and a gel mobility shift assay. As a result, all of the PNA-tolane derivatives stabilized duplex formation to the matched target sequence without inducing mismatch target binding. Among the different PNA-tolane derivatives, PNA that was modified with a naphthyl-type tolane could efficiently discriminate a mismatched base pair and be utilized for the detection of resistance to neuraminidase inhibitors of the influenza A/H1N1 virus. Therefore, our molecular tool can be used to discriminate single nucleotide polymorphisms that are related to drug resistance in pathogens.

Keywords: drug resistance; influenza virus; peptide nucleic acid; single nucleotide polymorphism; tolane.

MeSH terms

  • DNA / chemistry
  • DNA, Single-Stranded / chemistry
  • Drug Resistance*
  • Humans
  • Molecular Diagnostic Techniques*
  • Molecular Structure
  • Nucleic Acid Conformation
  • Peptide Nucleic Acids* / chemical synthesis
  • Peptide Nucleic Acids* / chemistry
  • Polymorphism, Single Nucleotide*
  • RNA / chemistry
  • Structure-Activity Relationship
  • Thermodynamics

Substances

  • DNA, Single-Stranded
  • Peptide Nucleic Acids
  • RNA
  • DNA