A number of small molecules bind directly and selectively to DNA, acting as chemotherapeutic agents by inhibiting replication, transcription or topoisomerase activity. Two common binding modes for these small molecules are intercalation or groove-binding. Intercalation results from insertion of a planar aromatic substituent between DNA base pairs, with concomitant unwinding and lengthening of the DNA helix. Groove binding, in contrast, does not perturb the duplex structure to any great extent. Groove-binders are typically crescent-shaped, and fit snugly into the minor groove with little distortion of the DNA structure. Recent calorimetric studies have determined the enthalpic and entropic contributions to the DNA binding of representative DNA binding compounds. Analysis of such thermodynamic data culled from the literature reveals distinctive thermodynamic signatures for groove-binding and intercalating compounds. Plots of the binding enthalpy (DeltaH) against binding entropy (-TDeltaS) for 26 drug-DNA interactions reveal that groove-binding interactions are clustered in a region of the graph with favorable entropy contributions to the free energy, while intercalators are clustered in a region with unfavorable entropy but favorable enthalpy contributions. Groove-binding is predominantly entropically driven, while intercalation in enthalpically driven. The molecular basis of the contrasting thermodynamic signatures for the two binding modes is by no means clear, but the pattern should be of use in categorizing new DNA binding agents.