A quantitative assay for the non-covalent association between apolipoprotein(a) and apolipoprotein B: an alternative measure of Lp(a) assembly

Increasing evidence suggests that the assembly of lipoprotein(a) (Lp(a)) proceeds in two steps. In the first step, non-covalent interactionsbetween apolipoprotein(a) (apo(a)) and apolipoprotein B(apoB) of low density lipo- protein (LDL) form a dissociable apo(a):LDL complex. In the second step, a covalent disulfide linkage forms the sta- ble Lp(a) particle. Several methods are currently used to study the assembly of Lp(a), however, these methods are laborious, time-consuming, and not suitable for a high throughput screening. We report here the development of a rapid and simple assay based on the binding of labeled LDL to a Lp(a)/apo(a) substrate which is immobilized on the surface of a microtiter plate. Quantification of bound LDL provides a measure of the extent of complex formation. La- beled LDL bound to both Lp(a) and apo(a) substrates with similar affinity. Plasma lipoproteins containing apoB as well as free apo(a) were capable of competing with LDL bind- ing. The binding of LDL to Lp(a)/apo(a) was inhibited by L-proline and lysine analogs, which are known to inhibit the non-covalent association between apo(a) and apoB. Using this method we have found that nicotinic acid and captoprilare able to inhibit the association of apo(a) with apoB. This method is compatible with automation and can be applied to a high throughput screeningof inhibitors of Lp(a) for- mation. —Dardik, B. N., C. D. Schwartzkopf, D. E. Stevens, and R. E. Chatelain. A quantitative assay for the non-covalent association between apolipoprotein(a) and apolipoprotein B: an alternative measure of Lp(a) assembly. J. Lipid Res. 2000. 41: 1013-1019.