Structural Characterization of Co·Bleomycin A2 Brown:  Free and Bound to d(CCAGGCCTGG)

A model for the structure of the aquated form of cobalt bleomycin (H 2 O-CoBLM or CoBLM A2 brown), free and bound to d(CCAGG C CTGG) 2 ( 1 ) is reported based on molecular modeling using the constraints obtained from 2D NMR studies. CoBLM A2 brown has a chiral organization of its ligands, including the axial primary amine of β-aminoalanine, identical to the hydroperoxide form of cobalt BLM (HOO-CoBLM or CoBLM A2 green). H 2 O-CoBLM forms a 1:1 complex with 1 with a K d of 2 × 10 - 6 M which is in slow exchange on the NMR time scale. The complex exhibits 44 intermolecular NOEs and 56 intramolecular NOEs within H 2 O-CoBLM itself. Molecular modeling reveals that H 2 O-CoBLM's mode of binding, basis for sequence specificity, and chemical specificity are almost identical to that previously reported for HOO-CoBLM (Wu, W.; Vanderwall, D. E.; Turner, C. J.; Kozarich, J. W.; Stubbe, J. J. Am. Chem. Soc. 1996, 118, 1281−1294). The bithiazole tail is inserted from the minor groove 3‘ to C 6 and the terminal thiazolium ring is well stacked between the bases G14 and G15, while the penultimate thiazolium ring is only partially stacked between the bases of C 6 and C7. The basis for sequence specificity involves hydrogen-bonding interactions between the 4-amino group and N3 of the pyrimidine of BLM and the N3 and 2-amino group of G5 of DNA, forming an unusual base triple. Molecular modeling further reveals that the oxygen of the axial H 2 O ligand is 2.7 Å from the 4‘ H of C 6, the site of cleavage of iron BLM and light-activated cleavage by HOO-CoBLM. Preliminary studies of H 2 O-CoBLM with d(CCAGTACTGG) ( 4 ) and d(GGAAGCTTCC) ( 2 ) are also reported.