A NEW CLASS OF ARTIFICIAL NUCLEASES THAT RECOGNIZE AND CLEAVE APURINIC SITES IN DNA WITH GREAT SELECTIVITY AND EFFICIENCY

A series of tailor-made molecules, 1 and 4-7, have been prepared to re cognize and cleave DNA at apurinic sites. These molecules incorporate in their structure different units designed for specific functions: (1 ) an intercalator for DNA binding, (2) a nucleic base for abasic site recognition, and (3) a linker endowed with both a binding function and a cleavage function (Scheme II). The constituent units were varied su ccessively in the series of molecules to get insight into their mode o f action and prepare more active compounds. H-1 NMR spectroscopy revea ls the absence of intramolecular ring-ring stacking interactions in wa ter between the base and the intercalator in all molecules 1 and 4-7. All bind to calf thymus DNA with binding constants ranging from 10(4) to 10(6) M-1. Their nuclease activity was estimated by measuring their ability to induce single strand breaks in depurinated pBR 322 plasmid DNA. The most efficient molecule, 5, exhibits high recognition select ivity and cleavage efficiency: at nanomolar concentrations, 5 recogniz es and cleaves the abasic lesion present in a DNA molecule containing an average of 1.8 apurinic sites in its 4 362 base pairs sequence. Mol ecule 5 exhibits higher cleaving efficiency than the reported tripepti de Lys-Trp-Lys: 10(-8) M concentrations of the former (5) lead to clea vage ratios comparable to those observed for the latter used as 10(-3) M concentration. This enzyme mimic 5 can be used advantageously as a substitute to the natural nuclease for in vitro cleavage of depurinate d DNA.