Sy. Breusegem et al., Experimental precedent for the need to involve the primary hydration layerof DNA in lead drug design, J MED CHEM, 44(16), 2001, pp. 2503-2506
The increase in fluorescence on binding of m-phenyl substituted hydroxy der
ivatives of Hoechst 33258 with poly[d(A-T)], d(CGCGAATTCGCG)(2), and with t
he corresponding T-4-looped 28-mer AATT hairpin was used to monitor binding
by equilibrium titrations and stopped-flow kinetics. Replacing the p-OH su
bstituent of Hoechst 33258 (association constant K-a = 5.2 x 10(8) M-1 for
28-mer hairpin) by m-OH increases the AATT site binding energy by 1.1 kcal
mol(-1), K-a = 3.8 x 10(9) M-1. Addition of a second m-hydroxy group (bis-m
-OH Hoechst) further strengthens binding, giving K-a = 1.9 x 10(10) M-1, an
d the binding energy increases by about 2.1 kcal mol(-1) compared to p-OH H
oechst. The value of K-a determined at equilibrium equaled that determined
from the ratio of association and dissociation rate constants from stopped-
flow studies. The increase in affinity of the monohydroxy Hoechst analogue
(m-OH) may originate from water-mediated hydrogen bonding with the minor gr
oove. The further increase in affinity of the bis-m-OH derivative (whose se
cond m-OH group must be directed away from the DNA minor groove floor) may
arise from a hydrogen-bonded network of water molecules. The potential to i
ncrease binding strength through relayed water molecules is proposed as an
additional possible input for lead drug design at DNA targets.