Peptide nucleic acids (PNA) are mimics with normal bases connected to a pseudopeptide chain that obey Watson-Crick rules to form stable duplexes with itself and natural nucleic acids. This has focused attention on PNA as therapeutic or diagnostic reagents. Duplexes formed with PNA mirror some but not all properties of DNA. One fascinating aspect of PNA biochemistry is their reaction with enzymes. Here we show an enzyme reaction that operates effectively on a PNA͞DNA hybrid duplex. A DNA oligonucleotide containing a cis, syn-thymine [2؉2] dimer forms a stable duplex with PNA. The hybrid duplex is recognized by photolyase, and irradiation of the complex leads to the repair of the thymine dimer. This finding provides insight into the enzyme mechanism and provides a means for the selective repair of thymine photodimers.UV light damages nucleic acids primarily by causing dimerization of adjacent pyrimidines. This damage is repaired enzymatically by DNA photolyase, which binds selectively to cis, syn-cyclobutane dimers and, when this complex is activated by light, reforms the monomeric pyrimidines (1, 2). These reactions are outlined in Scheme 1 for thymines. Thymine photodimerization and its repair is an appealing system for examination of parallels in the properties of peptide nucleic acids (PNA), Scheme 2, and DNA. Photolyase is a siteselective, rather than a sequence-selective, enzyme. It binds to thymine dimers whether they are incorporated in superhelical, circular, linear, or single-stranded DNA. It appears that the only additional requirement for recognition by the enzyme is phosphate groups on the dimer-containing strand (3, 4). For these reasons, we suspected that photolyase might retain its ability to recognize and repair thymine dimers contained on the DNA strand of a DNA͞PNA hybrid duplex (5-8). We carried out a series of experiments to assess this possibility.
MATERIALS AND METHODSMelting Temperature (T m ) Measurements. Absorbance versus temperature curves were measured at 260 nm in 10 mM phosphate buffer at pH 7 for solutions containing 2 M of each strand of PNA or DNA shown in Scheme 3. The melting temperature, T m , is assigned as the peak of the first derivative plot.Gel Mobility Assay. The oligonucleotides DNA(1) and DNA(3) were labeled with 32 P at the 5Ј end by using standard techniques (9). Separate samples of the radiolabeled DNA (2,500 cpm) were mixed with complementary DNA(2) (5 M), PNA(1), or PNA(4) (5 M) in 10 l of 10 mM phosphate buffer at pH 7. The hybridization was carried out by heating to 90°C for 5 min and then cooling the solution to room temperature for 2 h. The samples were dried with a Speedvac at low heat for 2 h, and 5 l of nondenaturing loading buffer was added. The analysis was carried out by electrophoresis in a 20% polyacrylamide gel (29:1 acrylamide and bisacrylamide) followed by autoradiography. The gel was run with TBE buffer containing 89 mM Tris-borate and 1 mM EDTA, pH 8.