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Arginine-PNAs
monomers in the middle position of a PNA strand resulted in an excellent PNA probe (called the
“chiral box” PNA), with good affinity and excellent selectivity 10,11 . “Chiral box” PNAs have been
used in several applications for the discrimination of point mutations, obtaining excellent results in
assays based on mass spectrometry and electrophoresis 12,13 .
Unfortunately, it is difficult to apply of these molecules on surface devices, such as PNA microarrays,
since the immobilization of the probes on the surface usually exploits the reaction between the
terminal amino group of the PNAs and a reactive group on the surface 14 . The presence of the lysine
primary amino groups, on its side chain, would thus interfere with this reaction.
In this chapter a new class of chiral PNAs will be presented, mainly intended for surface
applications, such as the development of chiral PNA microarrays. The general idea was to
substitute the side chain of lysine with that of arginine, which also contains a positive charge
but does not bear a primary amino group. A PNA with one 2D,5L-Arg modified monomer in
the middle, to be compared with the analogous lysine-based PNAs, and a PNA with a series
of chiral monomers resulting in four adjacent arginine side chains have been synthesized, and
their binding properties towards complementary oligonucleotides have been studied.
3.2 Results and Discussion
3.2.1 Sequence Design
The need to develop probes able to perform DNA recognition on surface determined the
necessity to substitute the previously used lysine, in the development of chiral PNAs. In fact,
the strategy usually employed to link the PNAs to glass surfaces is based on the reaction
between a reactive group on the surface and the terminal amino group of the PNA 14 . The
presence of the amino group of the lysine side-chains would thus compete with the terminal
one in this reaction, making the PNA deposition less reproducible. For this reason, chiral
modifications based on the structure of arginine were chosen, since the guanidium groups are
much less nucleophilic than primary amines, but still bear a positive charge at physiological
pH.
Since it has been demonstrated that PNAs containing one residue with two lysine-derived side
chains having configurations 2D and 5L had a highly improved affinity and selectivity in the
recognition of complementary DNA sequences, a PNA was first designed bearing a 2D,5Larginine
monomer in the middle (PNA 1, Figure 3-2), in order to compare its binding abilities
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