Key Points
- To identify an endothelial 116-kDa non-proteoglycan binding partner for lipoprotein lipase and evaluate whether it facilitates enzyme binding to endothelial surfaces.
- Purified the 116-kDa heparin-releasable protein from endothelial extracts using heparin-agarose chromatography, followed by Lys-C protease digestion and microsequencing.
- Assessed lipoprotein lipase (LPL) binding to apoB fragments from transfected CHO cells and thrombin-digested low-density lipoproteins using ligand blotting.
- Quantified 125I-LPL binding to endothelial cells in the presence of domain-specific anti-apoB monoclonal antibodies (mAb3, mAb19, mAb47), soluble apoB fragments, and heparin.
- Microsequencing and Western blot analysis identified the 116-kDa binding factor as an amino-terminal fragment of human apolipoprotein B present on endothelial cell membranes.
- Monoclonal antibodies targeting epitopes near the amino terminus of apoB (mAb3 and mAb19) inhibited 125I-LPL cellular binding by 60% to 65%, compared with approximately 10% inhibition by the carboxyl-terminal antibody mAb47.
- Heparin treatment eliminated the inhibitory effect of mAb3 and mAb19 by removing the 116-kDa protein, while pre-incubation with an amino-terminal apoB fragment reduced LPL cellular binding.
Structured PICO
PPopulationEndothelial cells, apoB-transfected CHO cells, and low density lipoprotein
IInterventionMonoclonal antibodies against apoB (mAb3, mAb19) and NH2-terminal apoB fragments
CComparatorControl conditions (mAb47, untreated cells)
OOutcome125I-LPL binding to cellssurrogate
An amino-terminal fragment of apolipoprotein B on endothelial cell surfaces facilitates the binding of lipoprotein lipase, providing new insights into lipid metabolism and endothelial interactions.