Influence of vesicle surface composition on the interfacial binding of lecithin:cholesterol acyltransferase and apolipoprotein A-I

J Lipid Res. 1997 Jun;38(6):1094-102.

Abstract

Interfacial binding affinities and capacities of lecithin:cholesterol acyltransferase (LCAT) and apolipoprotein A-I (apoA-I) for surfaces of different phosphatidylcholine (PC) composition, cholesterol content, and apolipoprotein content were measured with a vesicle model system. Native polyacrylamide gel electrophoresis was used to separate free protein from vesicle-bound protein. ApoA-I was isolated from human plasma and radiolabeled with iodine, whereas radiolabeled LCAT was purified from the media of Chinese hamster ovary cells that were transfected with human LCAT cDNA and incubated in the presence of [35S] cysteine and methionine. Bound and free radiolabeled LCAT and apoA-I were quantified by phosphorimage analysis. ApoA-I binding was not influenced by cholesterol content (14 mole%) but was influenced by the PC fatty acyl composition of the vesicle. PC species containing long chain, polyunsaturated fatty acids (PUFA) in the sn-2 position resulted in increased binding affinity (Kd = 75-177 nM) but reduced capacity (0.1-0.3 apoA-I/ 1000 PC) in comparison to sn-1 palmitoyl, sn-2 oleoyl PC (POPC, 750 nM and 1.4 apoA-I/1000 PC). LCAT binding affinity to POPC (2190 nM) was stronger in the presence of cholesterol (530 nM), and LCAT binding capacity was reduced (2.63 and 0.6 molecules LCAT/1000 PC, respectively). In comparison to POPC, LCAT binding affinity to sn-1 palmitoyl, sn-2 arachidonyl PC was stronger (611 nM) and binding capacity was reduced (0.7 LCAT/1000 PC). LCAT binding affinity and capacity to sn-1 palmitoyl, sn-2 eicosapentaneoyl PC (2041 nM, and 2.5 LCAT/1000 PC) were similar to those observed for POPC. We conclude that vesicle surface PC fatty acyl composition and cholesterol content significantly influence LCAT and apoA-I interfacial binding and therefore may alter LCAT enzymatic activity.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Apolipoprotein A-I / isolation & purification
  • Apolipoprotein A-I / metabolism*
  • Binding, Competitive
  • CHO Cells
  • Cholesterol / metabolism
  • Cricetinae
  • Dose-Response Relationship, Drug
  • Electrophoresis, Polyacrylamide Gel
  • Humans
  • Liposomes / chemistry
  • Liposomes / metabolism
  • Phosphatidylcholine-Sterol O-Acyltransferase / biosynthesis
  • Phosphatidylcholine-Sterol O-Acyltransferase / chemistry
  • Phosphatidylcholine-Sterol O-Acyltransferase / genetics
  • Phosphatidylcholine-Sterol O-Acyltransferase / metabolism*
  • Phosphatidylcholines / chemistry*
  • Phosphatidylcholines / metabolism
  • Sulfur Radioisotopes
  • Surface Properties
  • Transfection

Substances

  • Apolipoprotein A-I
  • Liposomes
  • Phosphatidylcholines
  • Sulfur Radioisotopes
  • Cholesterol
  • Phosphatidylcholine-Sterol O-Acyltransferase