Pseudocomplementary PNAs as selective modifiers of protein activity on duplex DNA: The case of type IIs restriction enzymes

Abstract

This study evaluates the potential of pseudocomplementary peptide nucleic acids (pcPNAs) for sequence-specific modification of enzyme activity towards double-stranded DNA (dsDNA). To this end, we analyze the ability of pcPNA-dsDNA complexes to site-selectively interfere with the action of four type IIs restriction enzymes. We have found that pcPNA-dsDNA complexes exhibit a different degree of DNA protection against cleaving/nicking activity of various isoschizomeric endonucleases under investigation (Plel, Mlyl and N.BstNBI) depending on their type and mutual arrangement of PNA-binding and enzyme recognition/cleavage sites. We have also found that the pcPNA targeting to closely located Plel or Bbsl recognition sites on dsDNA generates in some cases the nicking activity of these DNA cutters. At the same time, Mlyl endonuclease, a Plel isoschizomer, does not exhibit any DNA nicking/cleavage activity, being completely blocked by the nearby pcPNA binding. Our results have general implications for effective pcPNA interference with the performance of DNA-processing proteins, thus being important for prospective applications of pcPNAs.