Characterization of a 95 kDa high affinity human high density lipoprotein-binding protein

A new human 95 kDa high density lipoprotein (HDL)-binding protein (HBP) cor responding to a high affinity HDL-binding site with K-d = 1.67 mug/mL and a capacity of 13.4 ng/mg was identified in human fetal hepatocytes. The HDL binding with the 95 kDa HBP plateaus at 2.5-5 mug/mL under reducing and non reducing conditions. The association of HDL3 with the 95 kDa HBP plateaued in 15-30 min while dissociation was complete in 30 min. HDL3, apoA-I, and a poA-II were recognized by the 95 kDa HBP while low density lipoproteins (LD L) and tetranitromethane-modified HDL were not. The 95 kDa HBP predominantl y resides on the surface of cells since trypsin treatment of HepG2 cells el iminated nearly 70% of HDL binding. All studied human cells and cell lines (HepG2, Caco-2, HeLa, fibroblasts, SKOV-3, PA-I) demonstrated the presence of the 95 kDa protein. Both RT-PCR and Western blotting for HB-2/ALCAM were negative in human fetal hepatocytes while Gp96/GRP94 was clearly different iated from the 95 kDa HBP by two-dimensional electrophoretic mobility. More over, deglycosylation of HepG2 membrane preparations did not affect either HDL binding to the 95 kDa HBP or its size, while in contrast it affected th e molecular weights of HB-2/ALCAM and SR-BI/CLA-1. We conclude that the 95 kDa HBP is a new HDL receptor candidate widely expressed in human cells and cell lines.