Direct demonstration in synthetic oligonucleotides that N,N '-bis(2-chloroethyl)-nitrosourea cross-links N-1 of deoxyguanosine to N-3 of deoxycytidine on opposite strands of duplex DNA

The sequence specificity and covalent structure of the lesion caused by the DNA interstrand cross-linking reaction of N,N'-bis(2-chloroethyl)nitrosour ea (BCNU) were investigated using synthetic oligonucleotides, The efficienc y of interstrand cross-linking was found to parallel the efficiency of mono adduct formation, preferring deoxyguanosine-deoxycytidine-rich duplexes and , particularly, runs of deoxyguanosine. No explicit sequence specificity wa s observed. Enzymatic digestion of purified, interstrand cross-linked DNA r eturned primarily the unmodified deoxynucleosides, along with 1-[N-3-deoxyc ytidyl]-2-[N-1-deoxyguanosyl]ethane. This substance was characterized by co mparison of its mass spectrum, high-pressure liquid chromatography retentio n time, and UV spectrum to an authentic standard prepared by chemical synth esis. These studies provide the first direct evidence that BCNU has no stro ng sequence preference for interstrand cross-linking and that substance 4, which has been previously isolated from BCNU-treated DNA, derives from alky lation on opposite strands of DNA, The lack of sequence preference and lesi on structure together suggest that one source of BCNU interstrand cross-lin ks is linkage of deoxyguanosine and deoxycytidine partners from a single bp .