The relationship between the effect of lysine analogues and salt on the conformation of lipoprotein(a).

Lipoprotein(a) [Lp(a)] exhibits many of the same properties as plasminogen, owing to a similar structural makeup from a composite of multiple kringle domains. Shared behavior includes induction of an expanded conformation by lysine analogues, inhibition of this effect, and creation of a compact conformation by NaCl. Here, we examine in detail the independent and mutual effects of NaCl and 6-aminohexanoic acid (6-AHA) on the structure of Lp(a) and the relationship between the binding of the two ligands. We find that NaCl promotes the compact conformation while binding to Lp(a) homogeneously. In the absence of salt, 6-AHA leads to the complete unfolding of Lp(a), a process that is accompanied by cooperative binding. Reversal of conformation and weakening of binding occurred when one ligand was added to Lp(a) in the presence of the other, suggesting competitive binding. High concentrations of NaCl completely reversed the expansion of Lp(a) in 100 mM 6-AHA, and high concentrations of 6-AHA unfolded Lp(a) in the presence of 100 mM NaCl, but only by 30% in the case of the 15 kringle IV Lp(a) studied. Induction of the compact form of Lp(a) appears to be an effect in common with all salts examined and cannot be attributed solely to the anion, as in the case of plasminogen. The results were summarized in terms of a model of Lp(a) depicting the conformational alterations of apo(a) caused by the binding of the two ligands. In the compact conformation in NaCl, apo(a) is apposed to the particle surface. The fully expanded form in 6-AHA results from release of both the variable and constant kringle domains. In the intermediate form in water and in a solution containing both NaCl and 6-AHA, only the variable domain is released from the particle surface.

Effect of phospholipase A2 digestion on the conformation and lysine/fibrinogen binding properties of human lipoprotein[a].

In vitro hydrolysis of human lipoprotein[a] (Lp[a]) by phospholipase A2 (PLA2) decreased the phosphatidylcholine (PC) content by 85%, but increased nonesterified fatty acids 3.2-fold and lysoPC 12.9-fold. PLA2-treated Lp[a] had a decreased molecular weight, increased density, and greater electronegativity on agarose gels. In solution, PLA2-Lp[a] was a monomer, and when assessed by sedimentation velocity it behaved like untreated Lp[a], in that it remained compact in NaCl solutions but assumed the extended form in the presence of 6-amino hexanoic acid, which was shown previously to have an affinity for the apo[a] lysine binding site II (LBS II) comprising kringles IV5-8. We interpreted our findings to indicate that PLA2 digestion had no effect on the reactivity of this site. This conclusion was supported by the results obtained from lysine Sepharose and fibrinogen binding experiments, in the presence and absence of Tween 20, showing that phospholipolysis had no effect on the reactivity of the LBS-II domain. A comparable binding behavior was also exhibited by the free apo[a] derived from each of the two forms of Lp[a]. We did observe a small increase in affinity of PLA2-Lp[a] to lysine Sepharose and attributed it to changes in reactivity of the LBS I domain (kringle IV10) induced by phospholipolysis. In conclusion, the extensive modification of Lp[a] caused by PLA2 digestion had no significant influence on the reactivity of LBS II, which is the domain involved in the binding of apo[a] to fibrinogen and apoB-100. These results also suggest that phospholipids do not play an important role in these interactions.