Induced Fit Conformational Changes of a 'Reversed Amidine' Heterocycle: Optimized Interactions in a DNA Minor Groove Complex

To better understand the molecular basis for recognition of the DNA minor groove by heterocyclic cations, a series of "reversed amidine" substituted heterocycles has been prepared. Amidine derivatives for targeting the minor groove have the amidine carbon linked to a central heterocyclic system, whereas in the reverse orientation, an amidine nitrogen provides the link. The reverse system has a larger dihedral angle as well as a modified spatial relationship with the groove relative to amidines. Because of the large dihedral, the reversed amidines should have reduced binding to DNA relative to similar amidines. Such a reduction is observed in footprinting, circular dichroism (CD), biosensor-surface plasmon resonance (SPR), and isothermal titration calorimetric (ITC) experiments with DB613, which has a central phenyl-furan-phenyl heterocyclic system. The reduction is not seen when a pyrrole (DB884) is substituted for the furan. Analysis of a number of derivatives defines the pyrrole and a terminal phenyl substituent on the reversed amidine groups as critical components in the strong binding of DB884. ITC and SPR comparisons showed that the better binding of DB884 was due to a more favorable binding enthalpy and that it had exceptionally slow dissociation from DNA. Crystallographic analysis of DB884 bound to an AATT site shows that the compound was bound in the minor groove in a 1:1 complex as suggested by CD solution studies. Surprisingly, unlike the amidine derivative, the pyrrole -NH of DB884 formed an H-bond with a central T of the AATT site and this accounts for the enthalpy-driven strong binding. The structural results and molecular modeling studies provide an explanation for the differences in binding affinities for related amidine and reversed amidine analogues.