Nuclease resistance and protein recognition properties of DNA and hybrid PNA-DNA four-way junctions

Abstract

Nucleic acids possess unique biochemical features that make them ideal candidates to inhibit “difficult to target” proteins. The limited stability of nucleic acids in vivo presents a major obstacle to their development as drugs. Here, immobile four-way junctions (4WJs) are used to target the DNA-binding cytokine, High Mobility Group B1. Hybrid 4WJs composed of DNA and peptide nucleic acids (PNA) are investigated. PNA possess enhanced nuclease stability vs. DNA. 4WJs are incubated with Exonuclease III and DNase I. The nuclease assays show that 4WJs containing multiple PNAs possess significantly higher stability. Circular dichroism assays are used to probe the groove topology of 4WJs with the minor groove binder, DAPI. The CD data indicates that multi-PNA 4WJs possess altered minor groove dimensions that reduces DAPI binding affinity. Logic suggests that the minor groove of multi-PNA hybrids possess significant perturbations to the topology and local electrostatic environment that prevents proper binding/recognition by nucleases and thus enhances stability.