The solution structure of bis(phenazine-1-carboxamide)-DNA complexes: MLN 944 binding corrected and extended

ABSTRACT MLN 944 is a bisintercalating DNA‐binding antitumor agent known to be a template inhibitor of transcription. Previous 1H NMR studies of its d(ATGCAT)2 complex concluded that its phenazine chromophores are protonated. However, we find that this is not so, which has important consequences for the charged state of the ligand, for the orientation of its 1‐carboxamide group in the complex, and for the details of the interaction of its protonated interchromophore linker with the DNA base pairs. Here, we report a corrected solution structure of the MLN 944‐d(ATGCAT)2 complex, and extend the study to complexes with d(TATGCATA)2, and d(TACGCGTA)2, using a variety of 1H and 31P NMR methods and molecular dynamics simulations employing the AMBER 12 force field. We find that for all three complexes MLN 944 binds as a dication, in which the chromophores are uncharged, in the DNA major groove spanning the central 2 GC base pairs in a manner that maintains the dyad symmetry of the DNA. The carboxamide group lies in the plane of the chromophore, its NH making hydrogen bonding interactions with the phenazine N10 nitrogen, and the protonated linkers form hydrogen bonds with the O6 atom of guanine. The dynamics simulations reveal extensive solvent interactions involving the linker amines, the carboxamide group, and the DNA bases. © 2014 Wiley Periodicals, Inc. Biopolymers 101: 1099–1113, 2014.