Related Experiment Videos
Clusterin (complement lysis inhibitor) forms a high density lipoprotein complex with apolipoprotein A-I in human
D E Jenne1, B Lowin, M C Peitsch
1Institut of Biochemistry, University of Lausanne, Epalinges-sur-Lausanne, Switzerland.
Insights
Clusterin, a complement inhibitor, circulates in plasma bound to apolipoprotein A-I (apoA-I) within high-density lipoproteins (HDL). This complex may regulate both complement and lipid transport.
Area of Science:
- Biochemistry
- Immunology
- Lipidology
Background:
- Clusterin, also known as human complement lysis inhibitor (CLI), is a known inhibitor of the terminal complement cascade.
- The native form and plasma interactions of clusterin are not fully elucidated.
Purpose of the Study:
- To identify and characterize protein components that co-purify with clusterin from human plasma.
- To investigate the functional relationship between clusterin and its associated plasma proteins, particularly concerning lipid transport.
Main Methods:
- Affinity chromatography using anti-clusterin antibodies.
- Immunoblotting and amino acid sequencing for protein identification.
- Lipid analysis of isolated complexes.
- Electrophoretic techniques (free flow isotachophoresis) and density ultracentrifugation for complex characterization.
Main Results:
- Apolipoprotein A-I (apoA-I) was identified as a 28-kDa protein co-purifying with clusterin.
- Clusterin binds to delipidated apoA-I and high-density lipoproteins (HDL).
- The isolated apoA-I-clusterin complex contains lipids, primarily cholesterol and phospholipids, and is found in HDL fractions.
Conclusions:
- Clusterin circulates in human plasma as a high-density lipoprotein (HDL) complex with apolipoprotein A-I (apoA-I).
- This complex may function as both a complement cascade inhibitor and a regulator of lipid transport and redistribution.
Abstract:
Clusterin/human complement lysis inhibitor (CLI) is incorporated stoichiometrically into the soluble terminal complement complex and inhibits the cytolytic reaction of purified complement components C5b-9 in vitro. Using an anti-clusterin affinity column, we found that an additional protein component with a molecular mass of 28-kDa co-purifies with clusterin from human plasma. We show by immunoblotting and amino acid sequencing that this component is apolipoprotein A-I (apoA-I). By using physiological salt buffers containing 0.5% Triton X-100, apoA-I is completely dissociated from clusterin bound to the antibody column. Free clusterin immobilized on the antibody-Sepharose selectively retains apoA-I from total human plasma. Delipidated apoA-I and to a lesser extent ultracentrifugation-purified high density lipoproteins (HDL) adsorbed to nitrocellulose also have a binding affinity for purified clusterin devoid of apoA-I. The isolated apoA-I-clusterin complex contains approximately 22% (w/w) lipids which are composed of 54% (mole/mol) total cholesterol (molar ratio of unesterified/esterified cholesterol, 0.58), 42% phospholipids, and 4% triglycerides. In agreement with the low lipid content, apoA-I-clusterin complexes are detected only in trace amounts in HDL fractions prepared by density ultracentrifugation. In free flow isotachophoresis, the purified apoA-I-clusterin complex has the same mobility as the native clusterin complex in human plasma and is found in the slow-migrating HDL fraction of fasting plasma. Our data indicate that clusterin circulates in plasma as a HDL complex, which may serve not only as an inhibitor of the lytic terminal complement cascade, but also as a regulator of lipid transport and local lipid redistribution.