Synthesis and Characterization of Conformationally Preorganized, (R)-Diethylene Glycol-Containing γ-Peptide Nucleic Acids with Superior Hybridization Properties and Water Solubility

Abstract

Developed in the early 1990's, PNA has emerged as a promising class of nucleic acid mimic because of its strong binding affinity and sequence selectivity towards DNA and RNA, and resistance to enzymatic degradation by proteases and nucleases; however, the main drawbacks, as compared to other classes of oligonucleotides, are water solubility and biocompatibility. Herein we show that installation of a relatively small, hydrophilic (R)-diethylene glycol (`miniPEG') unit at the γ-backbone transforms a randomly-folded PNA into a right-handed helix. Synthesis of optically pure R-MPγPNA monomers is described, which can be accomplished in a few simple steps from a commercially available and relatively cheap Boc-L-serine. Once synthesized, R-MPγPNA oligomers are preorganized into a right-handed helix and hybridize to DNA and RNA with greater affinity and sequence selectivity, and are more water soluble and less aggregating than the parental PNA oligomers. The results presented herein have important implications for the future design and application of PNA in biology, biotechnology and medicine, as well as in other disciplines including drug discovery and molecular engineering.

Purification and assembly of thermostable Cy5 labeled γ-PNAs into a 3D DNA nanocage

“…This is similar to the 2x excess of PNApeptide required for quantitative binding to the same DNA nanocage. 22 Some PNA sequences show mild aggregation at concentrations as low as 1 mM, 24 which could prevent binding of some PNA to the DNA nanocage. The larger excess required for g-PNA1 may be due to its reduced solubility compared to g-PNA3.…”

“…The assembly process was rapid and could be performed at or below room temperature, providing mild conditions that preserved the function of the attached polypeptides. Substitutions to the gamma position on the PNA backbone (see Figure S2) have been shown to increase the solubility and thermal stability of the PNA strand, 24 as well as provide a handle to introduce functional groups without affecting its binding to a complementary strand. 25 Functionalizing PNA at the gamma position would also allow a peptide to be connected anywhere along the backbone or even at multiple locations to form stable cyclic PNA-peptides.…”

Pyrrolidinyl PNA with α/β-Dipeptide Backbone: From Development to Applications

“…16 Also, preorganization resulting from a simple substitution at the γ-position in aegPNA significantly improves its binding properties. 17 Various substituents can be placed at this position to improve water solubility 18 or cell penetration 19,20 or to incorporate other functions. 21,22 Because of its straightforward access via proline derivatives having well-defined stereochemistries, the pyrrolidine ring has been extensively used as a constraint element in the design of new PNA structures ( Figure 3).…”

Artificial Nucleic Acid Probes and Their Applications in Clinical Microbiology

“…In essence, this PNA structure, termed gPNA, is the first PNA oligomer to have sequenceunrestricted invasion capabilities while maintaining single base-pair specificity (Chenna et al, 2008;He, Rapireddy, Bahal, Sahu, & Ly, 2009). In addition, in order to address the issue of solubility using gPNAs, Sahu et al demonstrated that by incorporating, at the g-position of the backbone a diethylene glycol moiety, this results in a significant increase in solubility while not adversely affecting affinity or sequence specificity (Bahal, Sahu, Rapireddy, Lee, & Ly, 2012;Sahu et al, 2011).…”