Limited proteolysis of high density lipoprotein abolishes its interaction with cell-surface binding sites that promote cholesterol efflux

Armando J. Mendez, John F. Oram

Research output: Contribution to journalArticle

32 Scopus citations

Abstract

High-density lipoprotein (HDL) components remove cholesterol from cells by two independent mechanisms. Whereas HDL phospholipids pick up cholesterol that desorbs from the plasma membranes, HDL apolipoproteins appear to interact with cell-surface binding sites that target for removal pools of cellular cholesterol that feed into the cholesteryl ester cycle. Here we show that mild trypsin treatment of HDL almost completely abolishes this apolipoprotein-mediated cholesterol removal process. When HDL was treated with trypsin for various periods of time and then incubated with cholesterol-loaded fibroblasts, treatment for only 5 min reduced the ability of HDL to remove excess cholesterol from cellular pools that were accessible to esterification by the enzyme acyl CoA:cholesterol acyltransferase. This mild treatment digested less than 20% of HDL apolipoproteins and did not alter the lipid composition, size distribution, or electrophoretic mobility of the particles. Trypsin treatment of HDL for up to 1 h caused no further reduction in its ability to remove cellular cholesterol despite a greater than 2-fold increase in apolipoprotein digestion. Trypsin treatment of HDL also reduced its ability to deplete the cholesteryl ester content of sterol-laden macrophages. Promotion of cholesterol efflux from the plasma membrane by HDL phospholipids was unaffected by even extensive proteolysis. In parallel to the loss of cholesterol transport-stimulating activity, trypsin treatment of HDL for only 5 min nearly abolished its interaction with high-affinity binding sites on cholesterol-loaded fibroblasts. Reconstitution of trypsin-modified HDL with isolated apo A-I or apo A-II restored the cholesterol transport-stimulating activity of the particles. Thus a minor trypsin-labile fraction of HDL apolipoproteins is almost exclusively responsible for the apolipoprotein-dependent component of cholesterol efflux mediated by HDL particles.

Original languageEnglish (US)
Pages (from-to)285-299
Number of pages15
JournalBiochimica et Biophysica Acta - Lipids and Lipid Metabolism
Volume1346
Issue number3
DOIs
StatePublished - Jun 22 1997
Externally publishedYes

Keywords

  • Apolipoprotein
  • Cell-surface binding site
  • Cholesterol efflux
  • HDL

ASJC Scopus subject areas

  • Biochemistry
  • Endocrinology
  • Biophysics

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