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Albany 2001

category image Biomolecular
Stereodynamics
SUNY at Albany
June 19-23, 2001

Complexing of Duplex DNA with a Pair of Pseudocomplementary PNAs

Pseudocomplementary peptide nucleic acids (pcPNAs) have recently been introduced for sequence-unrestricted targeting of double-stranded DNA (dsDNA) (1). Along with ordinary guanine and cytosine, these modified PNAs carry 2,6-diaminopurine (D) and 2-thiouracil (sU) instead of adenine and thymine, respectively. The D and sU nucleobases hinder association of two pcPNAs while allowing a stable Watson-Crick-type pairing of pcPNAs with the DNA counterparts. It has been shown that appropriate pairs of pcPNAs hybridize efficiently to designated dsDNA targets containing all four nucleobases in a sequence-specific manner. The formation of pcPNA/DNA complex impedes the action of DNA processing enzymes (2). It has been proposed that pcPNA binding to designated dsDNA sites proceeds via a novel type of interaction ? double-duplex invasion (1,2).

To elucidate the mechanism of this mode of stand invasion, we have embarked on thorough kinetics studies of pcPNAs binding to dsDNA at various temperatures using the gel-shift assay. In parallel, the possibility of association of pseudocomplementary PNA oligomers with each other has been assayed by mixing curves (Job plots) and thermal denaturation experiments. Our data shed a new light on the mechanism of double-duplex invasion of pcPNA into duplex DNA (3).

References and Footnotes

  1. Lohse, J., Dahl, O. & Nielsen, P.E. (1999) Double duplex invasion by peptide nucleic acid: a general principle for sequence-specific targeting of double-stranded DNA, Proc. Natl. Acad. Sci. USA 96, 11804-11808.
  2. Izvolsky, K.I., Demidov, V.V., Nielsen, P.E. & Frank-Kamenetskii, M.D. (2000) Sequence-specific protection of duplex DNA against restriction and methylation enzymes by pseudocomplementary PNAs, Biochemistry 39, 10908-10913.
  3. This work was supported by a HFSP fellowship to E.P.

Ekaterina Protozanova, Vadim V. Demidov and Maxim D. Frank-Kamenetskii

Center for Advanced Biotechnology, Boston University, 36 Cummington Street, Boston, MA 02215, USA Phone: (617) 353-8492; Fax: (617) 353-8501; email: ekp@bu.edu