Thrombin, the ultimate proteinase of the coagulation cascade, is an attractive target for the treatment of a variety of cardiovascular diseases. Previously, a series of novel thrombin inhibitors, discovered by employing a powerful and new computer-assisted multiparameter optimization process (CADDIS), have been synthesized. We have now crystallized the complex of human alpha-thrombin with the most potent of these inhibitors, 8-5 (K(i)=3 nM), and have determined its 2.3A X-ray crystal structure. The Fourier map displayed clear electron density for the inhibitor. The central part of the inhibitor binds in an improved melagatran-like mode, while the structure identifies a d-tyrosine as P1 residue which forms a charged hydrogen bond with Asp 189 of thrombin. This is the first crystal structure of a thrombin-inhibitor complex, where an uncharged inhibitor residue makes hydrogen bonds within the thrombin S1 pocket. Additionally, novel favourable intermolecular hydrogen bonds of the inhibitor with the thrombin backbone become possible due to the d-configuration of the P1 residue. Two flanking voluminous side chains increase the strength of the subjacent hydrogen bonding system by shielding it from the bulk solvent.