Studies on the Lysine-Binding Sites of Human Plasminogen. The Effect of Ligand Structure on the Binding of Lysine Analogs to Plasminogen

Abstract

A method was described for measuring relative binding constants of lysine and its analogs to plasminogen and plasminogen kringle fragments. Plasminogen or kringle fragments adsorbed to lysine-Sepharose were eluted with increasing concentrations of lysine or other ligands, the concentration of ligand required to elute 50% of the protein being taken as a measure of the binding constant. The method is simple and was not dependent on monitoring conformational changes. Earlier reports were confirmed that the best ligands for the lysine binding sites of plasminogen are .omega.-amino acids containing 5 or 6 carbons. Glu-plasminogen (the native form with N-terminal glutamic acid) and Lys-plasminogen (a degraded form with N-terminal lysine), as well as the H chain fragments, kringle 4 and kringle 1 + 2 + 3, have very similar properties with regard to binding specificity for .omega.-amino acids. For all species optimal binding was observed when the distance between the amino and carboxyl carbon was about 0.68 nm. The binding of ligands was decreased by the presence of polar atoms on the .alpha. and .beta. positions of the carbon chain of amino acids. Arginine bound relatively weakly at the lysine site and there did not appear to be a separate arginine binding site in plasminogen.