BIOCHEMICAL AND PHYSICOCHEMICAL PROPERTIES OF PHOSPHORODITHIOATE DNA

The biochemical and physicochemical properties of DNA oligomers contai ning phosphorodithioate linkages in various configurations were evalua ted. Duplex stability studies, which were carried out by thermal denat uration analysis with complementary unmodified DNA, indicated a highly cooperative process similar to completely unmodified duplexes. Oligom ers containing phosphorodithioate linkages were found to have reduced melting temperatures relative to unmodified duplexes, with the degree of T-m depression paralleling the percent phosphorodithioate compositi on of the oligomer. Relative to activation of RNase H, DNA oligomers c ontaining up to 50% phosphorodithioate linkages were able to direct RN ase H degradation with the same efficiency as unmodified DNA while tho se containing from 50 to 100% acted with somewhat reduced efficiency. At Limiting concentrations, an oligomer containing alternating phospho rodithioate and phosphate linkages was able to direct RNase H degradat ion of the target RNA in an extended incubation, while an unmodified o ligomer did not. The nuclease resistance of phosphorodithioate-contain ing oligomers was evaluated in HeLa cell nuclear and cytoplasmic extra cts, in human serum, and with nucleases SI and DNase I. Oligomers cont aining alternating phosphorodithioate and phosphate were highly resist ant to degradation in all systems. However, oligomers having more than one unmodified linkage separating phosphorodithioates were degraded r apidly by DNase I, while demonstrating stability to degradation in all other systems tested. These results indicate that phosphorodithioate- containing DNA oligomers are highly nuclease-resistant, are able to fo rm stable duplexes with complementary nucleic acid sequences, and effi ciently direct RNase H degradation of target RNA.