Abstract
Various aspects of lipoprotein lipase (LPL) metabolism, including cell surface binding, degradation, and enzymatic activity, were compared between Chinese hamster ovary (CHO) cells and two distinct proteoglycan-deficient CHO cell lines. The contribution of low density lipoprotein receptor-related protein in binding LPL was also analyzed by the use of a 39-kDa receptor-associated protein expressed as a glutathione S-transferase fusion protein (GST-RAP). Equilibrium binding data with 125I-LPL revealed the presence of a class of high affinity binding sites with a KD of 7.8 nM in CHO cells, whereas no high affinity binding was observed for proteoglycan-deficient cells. The high affinity binding of LPL in CHO cells appeared to be concentrated in cell surface projections and was not effectively inhibited by GST-RAP. Moreover, degradation of endogenous and exogenous LPL was significantly greater in control CHO cells than in proteoglycan-deficient cells. Degradation of LPL in CHO cells was not affected by GST-RAP, suggesting that proteoglycans and not low density lipoprotein receptor-related protein are responsible for the majority of binding and degradation of LPL in these cells. Our data also show that proteoglycan binding is not essential for the assembly of active LPL homodimers, although proteoglycan binding controls the distribution of LPL activity. Furthermore, LPL produced by CHO cells was more stable than LPL produced by proteoglycan-deficient cells.
Highlights
Heparan sulfate proteoglycans (HSPGs) act as cell surface receptors that mediate the binding of lipoprotein lipase (LPL) to both parenchymal [2, 3] and endothelial (4 –7) cell types, the fate of bound LPL differs dramatically between the cell types
Do HSPGs function in other aspects of LPL metabolism? For example, are HSPGs necessary for the secretion of newly synthesized LPL? Are HSPGs solely responsible for LPL binding to cell surfaces or do other proteins participate? Are HSPGs or other LPL-binding proteins responsible for the intracellular degradation of LPL? Does HSPG binding influence the enzymatic activity of LPL? In most systems, it is difficult to definitively assign all of these functions to HSPGs, since most experimental systems contain lipoprotein receptor-related protein (LRP), gp330, or apolipoprotein B
We have addressed the significance of the LPL-HSPG interaction on the fate of LPL by the use of wild-type and mutant Chinese hamster ovary (CHO)-K1 cells
Summary
HSPGs act as cell surface receptors that mediate the binding of LPL to both parenchymal [2, 3] and endothelial (4 –7) cell types, the fate of bound LPL differs dramatically between the cell types. A number of conflicting reports show that binding of LPL to heparin, a proteoglycan very similar to heparan sulfate, can stabilize, stimulate, or inhibit catalytic efficiency of the lipase depending on experimental conditions [13,14,15,16]. Three additional proteins have been implicated in the cell surface binding of LPL These proteins are the low density lipoprotein receptor-related protein (LRP) [17, 18], gp330, known as megalin [19], and the amino-terminal fragment of apolipoprotein B [20, 21]. We investigated the contribution of LRP in the binding of LPL to these cells
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