LACTOFERRIN BINDING TO HEPARAN-SULFATE PROTEOGLYCANS AND THE LDL RECEPTOR-RELATED PROTEIN - FURTHER EVIDENCE SUPPORTING THE IMPORTANCE OF DIRECT BINDING OF REMNANT LIPOPROTEINS TO HSPG

Bovine lactoferrin inhibits the clearance of remnant lipoproteins from the plasma and competes with the cell-surface binding of apolipoprotein (ape) E-enriched remnants. We established that lactoferrin inhibits remnant binding and uptake by interacting with both heparan sulfate proteoglycans (HSPG) and the low-density lipoprotein receptor-related protein (LRP). The binding of I-125-lactoferrin was inhibited 45% to 60% in HepG2 hepatocytes and wild-type Chinese hamster ovary (CHO) cells treated with heparinase to remove HSPG. In mutant CHO cells (pgsD-677) lacking HSPG, the level of I-125-lactoferrin binding was approximate to 50% that seen with wild-type CHO cells; thus, about one half of lactoferrin binding appears to be mediated through cell-surface HSPG. A significant fraction of the residual binding of the lactoferrin appears to be mediated through the LRP. The 39-kd protein known to bind to the LRP and to block ligand interaction inhibited I-125-lactoferrin degradation in wild-type CHO cells by 60% to 65%. The addition of the 39-kd protein plus heparinase treatment reduced the binding by 85% to 90% (this combination blocks direct interaction with both the LRP and HSPG). However, it was also shown that the 39-kd protein bound to HSPG and the LRP. Heparinase treatment of wild-type CHO cells decreased the binding of the I-125-39-kd protein by approximate to 40%, and the mutant CHO cells lacking HSPG bound half as much I-125-39-kd protein as wild-type CHO cells. These studies also helped to establish that most of the enhanced binding of apoE-enriched beta-very-low-density lipoproteins (beta-VLDL) was via HSPG and not as a direct interaction with the LRP in the absence of HSPG. Whereas apoE-enriched beta-VLDL at a high concentration inhibited approximate to 45% of I-125-lactoferrin binding to wild-type CHO cells, I-125-lactoferrin binding to mutant CHO cells lacking HSPG (apparently binding to the LRP) was not inhibited by apoE-enriched beta-VLDL, thus further suggesting that apoB-enriched beta-VLDL does not interact to a major extent directly with the LRP in the absence of HSPG.