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 is described for measuring relative binding constants of lysine and analogs of lysine to plasminogen and plasminogen 'kringle' fragments. Plasminogen or kringle fragments adsorbed to lysine-Sepharose are 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 is not dependent on monitoring conformational changes. We confirm earlier reports that the best ligands for the lysine binding sites of plasminogen are w-amino acids containing five or six carbons. We show further that both Glu-plasminogen (the native form with N-terminal glutamic acid) and Lys-plasminogen (a degraded form with N-terminal lysine), as well as the heavy chain fragments, kringle 4 and kringle 1 + 2 + 3, have very similar properties with regard to binding specificity for w-amino acids. For all species optimal binding is observed when the distance between the amino and carboxyl carbon is about 0.68 nm. The binding of ligands is decreased by the presence of polar atoms on the a and p positions of the carbon chain of amino acids.Arginine binds relatively weakly at the lysine site and there does not appear to be a separate arginine binding site in plasminogen.It has been known for over 20 years that amino acids such as 6-aminohexanoic acid have antifibrinolytic properties [l], one of which probably results from inhibition of the binding of plasminogen to fibrin [2]. Subsequent studies have shown that 6-aminohexanoic acid and lysine cause extensive conformational alterations in Glu-plasminogen [3,4] (and references in [4]); Deutsch and Mertz [5] utilized the strong affinity of plasminogen for lysine-Sepharose in the development of a convenient method for purification of plasminogen.

Characterization of the interaction of bovine plasmin with Streptococcus uberis

“…The displacement of the inhibition/elution curves in each case reflects the generally acknowledged affinity of these amino acids for the lysine binding sites of plasmin and plasminogen (Winn et al 1980). The data obtained for Strep.…”

Modification of apolipoprotein(a) lysine binding site reduces atherosclerosis in transgenic mice.

“…Major physiological substrates of the LBS of plasminogen kringles 1 and 4 are fibrin and cell surface proteins containing carboxy-terminal lysines (27,44). This association results in the juxtaposition of plasminogen and plasminogen activators on two-dimensional substrates, greatly accelerating the conversion of the zymogen to active plasmin, as well as protecting it from subsequent deactivation by antiplasmins.…”

1H‐NMR assignments and secondary structure of human plasminogen kringle 1

“…7 5 nm [23], is the one which binds best to PgrdKl, K4 and K5, albeit with different affinities (Ka =77 mM-', 21 mM-' and 10 d-', respectively) . The tPA also interacts with fibrin [26].…”