In naked duplex DNA, G-C and A-T Watson-Crick base pairs exist in dynamic equilibrium with their Hoogsteen counterparts. Here, we used nuclear magnetic resonance (NMR) relaxation dispersion and molecular dynamics (MD) simulations to examine how Watson-Crick/Hoogsteen dynamics are modulated upon recognition of duplex DNA by the bisintercalator echinomycin and monointercalator actinomycin D. In both cases, DNA recognition results in the quenching of Hoogsteen dynamics at base pairs involved in intermolecular base-specific hydrogen bonds. In the case of echinomycin, the Hoogsteen population increased 10-fold for base pairs flanking the chromophore most likely due to intermolecular stacking interactions, whereas actinomycin D minimally affected Hoogsteen dynamics at other sites. Modulation of Hoogsteen dynamics at binding interfaces may be a general phenomenon with important implications for DNA-ligand and DNA-protein recognition.