CDB25:0003350 CLU — LRP2
Experimentally validated in Human, Mixed species; Orthology-inferred in Human, Mouse, Rat, Frog, Zebrafish, Chicken, Macaque, Pig, Dog, Cow, Chimp, Horse, Marmoset, Sheep
Title
Journal:; Year Published:
Abstract
Evidence that clusterin has discrete chaperone and ligand binding sites.
Biochemistry, 2002; PubMed, Homo sapiens CLU — Mus Musculus Lrp2
ABSTRACT: Clusterin is the first identified extracellular mammalian chaperone and binds to a wide variety of partly unfolded, stressed proteins.Clusterin also binds to many different unstressed ligands including the cell surface receptor low-density lipoprotein receptor-related protein-2 (LRP-2). It is unknown whether clusterin binds to all of these many ligands via one or more binding sites. Furthermore, the region(s) of clusterin involved in these many binding interactions remain(s) to be identified. As part of an investigation of these issues, we expressed recombinant human clusterin in the yeast Pichia pastoris. The resultant protein had variable proteolytic truncations of the C-terminal region of the alpha-chain and the N-terminal region of the beta-chain. We compared the chaperone and ligand binding activities of this recombinant product with those of clusterin purified from human serum. We also tested whether the binding of clusterin to ligands could be inhibited by competitive binding with other clusterin ligands or by anti-clusterin monoclonal antibodies. Collectively, our results indicate that (i) clusterin has three independent classes of binding sites for LRP-2, stressed proteins, and unstressed ligands, respectively, and (ii) the binding sites for LRP-2 and stressed proteins are likely to be in parts of the molecule other than the C-terminal region of the alpha-chain or the N-terminal region of the beta-chain. It has been suggested that, in vivo, clusterin binds to toxic molecules in the extracellular environment and carries these to cells expressing LRP-2 for uptake and degradation. This hypothesis is supported by our demonstration that clusterin has discrete binding sites for LRP-2 and other (potentially toxic) molecules.
Identification of glycoprotein 330 as an endocytic receptor for apolipoprotein J/clusterin.
The Journal of biological chemistry, 1995; PubMed, Homo sapiens CLU — Homo sapiens LRP2
ABSTRACT: Glycoprotein 330 (gp330) is a member of a family of endocytic receptors related to the low density lipoprotein receptor. gp330 has previously been shown to bind a number of ligands in common with its family member, the low density lipoprotein receptor-related protein (LRP). To identify ligands specific for gp330 and relevant to its localization on epithelia such as in the mammary gland, gp330-Sepharose affinity chromatography was performed. As a result, a 70-kDa protein was selected from human milk and identified by protein sequencing to be apolipoprotein J/clusterin (apoJ). Solid-phase binding assays confirmed that gp330 bound to apoJ with high affinity (Kd = 14.2 nM). Similarly, gp330 bound to apoJ transferred to nitrocellulose after SDS-polyacrylamide gel electrophoresis. LRP, however, showed no binding to apoJ in either type of assay. The binding of gp330 to apoJ could be competitively inhibited with excess apoJ as well as with the gp330 ligands apolipoprotein E, lipoprotein lipase, and the receptor-associated protein, a 39-kDa protein that acts to antagonize binding of all known ligands for gp330 and LRP. Several cultured cell lines that express gp330 and ones that do not express the receptor were examined for their ability to bind and internalize 125I-apoJ. Only cells that expressed gp330 endocytosed and degraded radiolabeled apoJ. Furthermore, F9 cells treated with retinoic acid and dibutyryl cyclic AMP to increase expression levels of gp330 displayed an increased capacity to internalize and degrade apoJ. Cellular internalization and degradation of radiolabeled apoJ could be inhibited with unlabeled apoJ, receptor-associated protein, and gp330 antibodies. The results indicate that gp330 but not LRP can bind to apoJ in vitro and that gp330 expressed by cells can mediate apoJ endocytosis leading to lysosomal degradation.