Double Duplex Invasion of DNA by PNA
The expanded repertoire of PNA binding motifs compared to other nucleic acids increases the potential for its exploitation in biotechnical, diagnostic, and therapeutic applications. A major limitation in the use of both PNA and oligonucleotides to date has been the inability to target mixed-sequence duplex DNA. In an exciting recent development, a new PNA targeting strategy has been reported in which a single-stranded PNA can invade mixed-sequence duplex DNA producing a novel structure with two identical PNA:DNA duplexes in the invasion complex.
PNA containing alternating diaminopurine and 2-thiouracil bases cannot form a self-complementary duplex because of steric hindrance between these modified bases and is thus termed pseudocomplementary. It can however form a very stable duplex with a complementary DNA strand containing alternating unmodified adenine and thymine bases, and can invade duplex DNA of this sequence to produce two DNA:PNA duplexes. This double duplex invasion has been monitored by CD spectroscopy which shows that the PNA bases become involved in a helical structure upon interaction with DNA. The stoichiometry of the final complex is confirmed to be 1:1 and the binding rate increases with PNA concentration and temperature, and decrease with ionic strength. Under pseudo-first order conditions, Arrhenius analysis reveals an ionic strength independent activation enthalpy of ca. 80 kJ mol-1 which indicates that, as for normal PNA triplex-strand-invasion, DNA basepair opening is likely the rate-limiting step. Differences in the double duplex invasion of single and multiple sites will be discussed.
Eimer Tuite, Peter Nielsen,* and Bengt Norden
*Center for Biomolecular Recognition, Department of Medical Biochemistry & Genetics,