FIGURE 5.

Reduced binding of human LPL to cells expressing mutant versions of GPIHBP1 with specific cysteine-to-alanine mutations in the Ly6 domain. CHO pgsA-745 cells were electroporated with an S-protein-tagged wild-type GPIHBP1 construct or mutant constructs with specific cysteine-to-alanine mutations. 24 h after the electroporation, the cells were incubated with V5-tagged human LPL in the presence or absence of heparin (500 units/ml). After washing the cells six times, the cell extracts were prepared, and the level of LPL bound to the cells was assessed by Western blotting with a V5 tag-specific antibody. Simultaneously, the level of GPIHBP1 in cell extracts was assessed by Western blotting with an antibody against the S-protein tag. Actin was used as a loading control.