Thermal melting studies of alkyne- and ferrocene-containing PNA bioconjugates

Abstract

The preparation of new metal-containing Peptide Nucleic Acids (PNAs) is currently a field of research intensively studied for various purposes, e.g. DNA biosensors. The role played by the metal centre, notably on the stability of the PNA.DNA hybrid, is obviously crucial, but has not yet been fully investigated. In this work, UV-Vis spectroscopic measurements of solutions of DNA.PNA hybrids, whose 11/12-mer PNA oligomers contained either one or two alkyne- (1) or ferrocene-containing (2) PNA monomers, were carried out to determine the effect of these monomers on the thermal stability of the hybrids (PNA: H-Gly-X-gggtc-Y-agctt-X-Lys-NH2 with X = 1 or and Y = 1 or 2 or blank position). Supplementary CD spectroscopic measurements were performed to gain insight into the structures of the PNA.DNA duplexes formed. The effect of both modified monomers was found to depend on their actual positions within the PNA sequences. Insertions at the N- or C-termini of a PNA oligomer did not change the melting temperatures (T(m) values of about 72 degrees C) of the DNA.PNA hybrids significantly. Insertion of monomers 1 or 2 in the middle of a PNA sequence induced a substantial decrease in the T(m) of the hybrids (by about 23 degrees C) when bound to the same DNA oligomer. Interestingly, it was found that the type of modification, namely alkyne or ferrocene, did not significantly influence the T(m) values in these cases. However, the thermal stability of hybrids with the DNA oligomers containing one to four additional thymines and the PNA oligomers containing the ferrocene moiety in its middle, varied significantly with the number of thymines added compared to its alkyne analogues (DeltaT(m) up to -13 degrees C). The presence of the ferrocene moiety induced a significant decrease in thermal stability of the hybrids, probably due to its bulkiness. In order to assess the effect of PNA backbone rigidity on the stability of DNA.PNA hybrids, PNA oligomers with an internal amino acid, propargylglycine (Pgl) or the dipeptide glycine-propargylglycine (Gly-Pgl), were synthesised. It was assumed that the orientation of the alkyne moiety in the Pgl-containing PNA sequence is not identical to an alkyne-containing PNA sequence, as a significantly higher T(m) value (DeltaT(m) = +10 degrees C) was measured. It is anticipated that the alkyne moiety in Pgl is not facing the DNA base and therefore does not disturb as much the neighbouring nucleobases and base-stacking of the complementary DNA, in contrast to the alkyne moiety of 1. Interestingly, no significant differences in the thermal stability of the hybrids was observed between Pgl-containing and dipeptide-containing PNA oligomers, although the former contracts the PNA backbone by three atoms.

Metal-containing peptide nucleic acid conjugates

“…The authors suggested that this difference could be explained by the different geometric preferences of Zn 2+ . 152 It was demonstrated that modification with alkyne-or ferrocene-containing monomers at the N-or C-terminus of the PNA sequence did not significantly alter the thermal stability of DNA•PNA hybrids. These hybrids were considered to be completely complementary due to their similar melting temperatures.…”

“…Loop formation in different PNA•DNA hybrids. 152 Adapted from reference. 152 Mokhir, Krämer and coworkers proposed to control the PNA binding to oligonucleotides with bioavailable metal ions.…”

“…152 Adapted from reference. 152 Mokhir, Krämer and coworkers proposed to control the PNA binding to oligonucleotides with bioavailable metal ions. Indeed, since the intracellular concentration of free transition metal ions is dependent on the cell type, 154 metal-responsive PNA•DNA/RNA binding could be employed for selective suppression of gene expression in these specific cells.…”

Chemical Biology: What is Its Role in Drug Discovery?

“…GNA containing fluorescent nucleotides have also been prepared (Hui Zhou et al, 2009). A novel nucleobase known as PPT has been synthesised and incorporated into PNA (Hirano et al, 2009) and interesting studies on novel metal-containing PNAs have been reported (Sosniak et al, 2009). The introduction of a disulfide bond into the main chain of PNA enabled the facile removal of the PNA from its duplex and invasion complexes with DNA (Aiba & Komiyama, 2009).…”

Specific uptake and interactions of peptide nucleic acid derivatives with biomimetic membranes

“…23-24, 36 70 In general terms, this article represents a significant addition to the growing field of research focussing on metal-containing PNAs. 13 Although much attention has been focussed on the preparation of such constructs [37][38][39][40][41] and only recently have papers appeared which report the applications of non-radioactive metal 75 bioconjugates in living cells. [14][15][42][43][44] It is therefore important to understand the mechanism of uptake of these nucleotide analogues.…”