SOLUTION STRUCTURE OF THE EPSILON-AMINOHEXANOIC ACID COMPLEX OF HUMANPLASMINOGEN KRINGLE-1

The solution structure of the human plasminogen kringle 1 domain compl exed to the antifibrinolytic drug 6-aminohexanoic acid (epsilon Ahx) w as obtained on the basis of H-1-NMR spectroscopic data and dynamical s imulated annealing calculations. Two sets of structures were derived s tarting from (a) random coil conformations and (b) the (mutated) cryst allographic structure of the homologous prothrombin kringle 1. The two sets display essentially the same backbone folding (pairwise root-mea n-square deviation, 0.15 nm) indicating that, regardless of the initia l structure, the data is sufficient to locate a conformation correspon ding to an essentially unique energy minimum. The conformations of res idues connected to prolines were localized to energetically preferred regions of the Ramachandran map. The Pro30 peptide bond is proposed to be cis. The ligand-binding site of the kringle 1 is a shallow cavity composed of Pro33, Phe36, Trp62, Tyr64, Tyr72 and Tyr74. Doubly charge d anionic and cationic centers configured by the side chains of Asp55 and Asp57, and Arg34 and Arg71, respectively, contribute to anchoring the zwitterionic EAhx molecule at the binding site. The ligand exhibit s closer contacts with the kringle anionic centers (approximate to 0.3 5 nm average O...H distance between the Asp55/Asp57 carboxylate and li gand amino groups) than with the cationic ones (approximate to 0.52 nm closest O...H distances between the ligand carboxylate and the Arg34/ Arg71 guanidino groups). The EAhx hydrocarbon chain rests flanked by P ro33, Tyr64, Tyr72 and Tyr74 on one side and Phe36 on the other. Dipol ar (Overhauser) connectivities indicate that the ligand aliphatic moie ty establishes close contacts with the Phe36 and Trp62 aromatic rings. The computed structure suggests that the EAhx molecule adopts a kinke d conformation when complexed to kringle 1, effectively shortening its dipole length to approximate to 0.65 nm.