Investigating the role of the N-terminus formamido group of distamycin

ORGN 688

Laura Westrate1, Hilary Mackay1, Binh Nguyen, chebkn@langate.gsu.edu2, W. David Wilson2, Jerome Kluza3, John Hartley3, Moses Lee, lee@hope.edu1, and Toni Brown1. (1) Department of Chemistry, Hope College, Natural Sciences Division, 35E. 12th. Street, Holland, MI 49422, (2) Department of Chemistry, Georgia State University, Atlanta, GA 30303, (3) Oncology, Royal Free & University College Medical School, 91 Riding House Street, London, W1W 7BS, United Kingdom
The N-terminus formamido-moiety (f-) of distamycin plays a crucial role in the binding of this naturally occurring polyamide in the minor groove of DNA: The f-group promotes a staggered dimeric binding motif, influences orientation specificity, and enhances the binding affinity. To further understand the function of the f-group, a series of polyamides with alternative N-terminus moieties [acetamido-(CH3-), amino- (NH2-) and trifluoroacetamido- (CF3-)] were synthesized. All compounds contained a heterocyclic core comprising of imidazole-pyrrole-imidazole (IPI). The binding characteristics of the above polyamides were evaluated using DNaseI footprinting and biophysical techniques. DNaseI footprinting of CH3-IPI demonstrated a clear selectivity for the cognate sequence (5'-ACGCGT-3' vs. 5'-AAATTT-3', 5'-ACCGGT-3' and 5'-A[T]A[G]T-3', where bases in square brackets denote a T/G mismatched base pair) and indicated that the acetamido N-terminus maintained orientation specificity in the 5' to 3' direction. Circular Dichroism demonstrated that CH3-IPI and NH2-IPI bound selectively to the cognate 5'-ACGCGT-3' sequence via the minor groove. SPR and ITC studies confirmed cognate selectivity and SPR provided a binding constant of 2.1 x 10(6) M-1 for CH3-IPI binding to 5'-ACGCGT-3, and studies are currently underway for NH2-IPI and CF3-IPI to further probe the selection of the f-group in nature.