PROTEOLYSIS AND FUSION OF LOW-DENSITY-LIPOPROTEIN PARTICLES STRENGTHEN THEIR BINDING TO HUMAN AORTIC PROTEOGLYCANS

Lipid droplets resembling those seen in the extracellular space of the arterial intima were generated in vitro when granule proteases of rat serosal mast cells degraded the apolipoprotein B-100 (apoB-100) compo nent of granule bound low density lipoprotein (LDL), and the particles fused on the granule surface (Paananen, K., and Kovanen, P. T. (1994) J. Biol. Chem. 269, 2023-2031). Moreover, the binding of the fused pa rticles to the heparin proteoglycan component of the granules was foun d to be strengthened. We have now treated LDL particles with alpha-chy motrypsin and examined the strength with which the proteolytically mod ified LDL binds to human aortic proteoglycans on an affinity column. W e found that chymotryptic degradation of the LDL particles triggered p article fusion. The higher the degree of proteolytic degradation, the higher were the degree of fusion and the strength of binding to the ao rtic proteoglycans. Separation of the proteolyzed particles by size ex clusion chromatography into two fractions, unfused and fused particles , and analysis of their binding strengths revealed that not only the f used but also the unfused proteolyzed particles bound more tightly to the proteoglycans than did the native LDL particles. To investigate th e mechanism underlying this increase in binding strength, we attached [C-13]dimethyl groups to the lysines and used NMR spectroscopy to quan tify the active lysine residues of apoB-100, which are thought to be l ocated in basic areas of apoB-100 and involved in binding of LDL to pr oteoglycans. Analysis of the C-13-labeled particles showed that, despi te loss of apoB-100 fragments from the particles, the number of active lysine residues in the unfused proteolyzed particles had not decrease d. In the fused proteolyzed particles, the number of active lysine res idues was markedly increased. Thus, proteolytic fusion appears to incr ease the number of basic domains of apoB-100, which would explain the observed increase in the strength of binding of the modified LDL parti cles to arterial proteoglycans. Since the fused particles resemble the small lipid droplets found in the atherosclerotic arterial intima, th is LDL modification offers a plausible mechanism for the focal accumul ation of lipid droplets in the extracellular proteoglycan matrix durin g atherogenesis.