ASSESSMENT OF THE ROLE OF THE BLEOMYCIN A(2) PYRIMIDOBLAMIC ACID C4 AMINO GROUP

The preparation and examination of 3 and 4 and their unnatural epimers 6 and 7 by following a new and alternative synthesis of 2 was conduct ed to assess the role of the pyrimidine C4 amine of bleomycin A(2) (1) and deglycobleomycin A(2) (2). The agent 3 bearing a pyrimidine C4 di methylamino substituent exhibited a substantially diminished DNA cleav age efficiency (10-15x) relative to 2 and the loss of the characterist ic 5'-GC/5'-GT cleavage selectivity. The agent 4 in which the pyrimidi ne C4 amino group was removed exhibited an even greater diminished DNA cleavage efficiency (30x) relative to that for 2. For this agent, the characteristic cleavage selectivity is either slightly or significant ly reduced depending on the assay conditions. Even in the instances wh ere it was not substantially altered, the ability to detect it require d a temperature of 4 versus 25-37 degrees C. This information and temp erature dependence suggest a reduced binding interaction and are consi stent with the participation of the pyrimidine C4 amine in one of two critical hydrogen bonds of a minor groove tripler-like recognition bet ween the metal binding domain of 1 and 2 and guanine at the 5'-GC/5'-G T cleavage sites responsible for the characteristic cleavage selectivi ty. These observations have further implications on the potential orig in of inherent 5'-GPy > 5'-APy cleavage selectivity of bleomycin A(2) itself. This cleavage preference of 5'-GPy > 5'-APy may be analogously attributed to a reduced binding affinity at the 5'-APy sites resultin g from one versus two tripler-like hydrogen bonds to adenine.