Sequence‐Unrestricted, Watson–Crick Recognition of Double Helical B‐DNA by (R)‐MiniPEG‐γPNAs

Development of general principles for designing molecules to bind sequence specifically to double-stranded DNA (dsDNA) has been a long-sought goal of bioorganic chemistry and molecular biology. Pursuit of this goal, in the past, has generally been focused on the minor and major grooves—in large part, because of the ease of accessibility of the chemical groups that reside on these external parts of the double helix and the precedence for their recognition in nature. It was long recognized that while Watson–Crick (W–C) base-pairing provides a more direct and specific means for establishing sequence-specific interactions with nucleic acid biopolymers, such as DNA and RNA, it would be difficult to do so with intact double helical DNA because of the preexisting base pairs. This effort has so far led to the development of several major classes of antigene molecules, with the likes of triplexforming oligonucleotides, minor-groove binding polyamides, and major-groove binding zinc-finger peptides. While they can be designed to bind sequence specifically to dsDNA, there are still remaining issues with sequence selection, specificity and/or target length that have not yet been completely resolved, 13, 17–19] although some progress has been made in recent years. Over the past two decades, peptide nucleic acids (PNAs), a particular class of nucleic acid mimics comprised of a pseudopeptide backbone (Scheme 1 A), have been shown to be capable of invading dsDNA. This finding is significant because, contrary to the traditional belief, it demonstrates that the DNA double helix is relatively dynamic at physiological temperatures, and that W–C base-pairing interactions can be established with intact dsDNA. Though promising as antigene reagents, because of the specificity of recognition and generality in sequence design, PNA binding is presently limited to mostly homopurine and homopyrimidine targets. Mixed-sequence PNAs have been shown to be capable of invading topologically constrained supercoiled plasmid DNA, conformationally perturbed regions of genomic DNA and duplex termini ; however, they are unable to invade the interior regions of double helical B-form DNA (B-DNA)—the most stable form of DNA double helix. “Tail-clamp” 31] and “doubleduplex invasion” strategies have subsequently been developed and have enabled mixed-sequence PNAs to invade BDNA, but they are not without limitations. Recently we showed that mixed-sequence PNAs, when preorganized into a right-handed helix by installing an (S)-Me stereogenic center at the g-backbone (Scheme 1 B) can invade BDNA. However, all of our studies so far have been limited to a few selected sequences due to the poor water solubility and propensity of these first-generation gPNAs to aggregate and adhere to surfaces and other macromolecules, including DNA, in a nonspecific manner. This problem is exacerbated Scheme 1. Chemical structures of: A) PNA, B) l-alanine-derived gPNA (gPNA), and C) (R)-MiniPEG-containing gPNA (gPNA). See ref. [36] for the synthesis of gPNA monomers; the methyl ether protecting group of miniPEG side-chain is removed in the final cleavage/deprotection step of oligomer synthesis.