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J. Biol. Chem., Vol. 282, Issue 25, 18602-18612, June 22, 2007
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Binding of Proprotein Convertase Subtilisin/Kexin Type 9 to Epidermal Growth Factor-like Repeat A of Low Density Lipoprotein Receptor Decreases Receptor Recycling and Increases Degradation*
1
2¶3
From the
Departments of Molecular Genetics and Internal Medicine, the Donald W. Reynolds Cardiovascular Clinical Research Center, and the ¶Howard Hughes Institute, University of Texas Southwestern Medical Center, Dallas, Texas 75390
Proprotein convertase subtilisin/kexin type 9 (PCSK9) promotes degradation of hepatic low density lipoprotein receptors (LDLR), the major route of clearance of circulating cholesterol. Gain-of-function mutations in PCSK9 cause hypercholesterolemia and premature atherosclerosis, whereas loss-of-function mutations result in hypocholesterolemia and protection from heart disease. Recombinant human PCSK9 binds the LDLR on the surface of cultured hepatocytes and promotes degradation of the receptor after internalization. Here we localized the site of binding of PCSK9 within the extracellular domain of the LDLR and determined the fate of the receptor after PCSK9 binding. Recombinant human PCSK9 interacted in a sequence-specific manner with the first epidermal growth factor-like repeat (EGF-A) in the EGF homology domain of the human LDLR. Similar binding specificity was observed between PCSK9 and purified EGF-A. Binding to EGF-A was calcium-dependent and increased dramatically with reduction in pH from 7 to 5.2. The addition of PCSK9, but not heat-inactivated PCSK9, to the medium of cultured hepatocytes resulted in redistribution of the receptor from the plasma membrane to lysosomes. These data are consistent with a model in which PCSK9 binding to EGF-A interferes with an acid-dependent conformational change required for receptor recycling. As a consequence, the LDLR is rerouted from the endosome to the lysosome where it is degraded.
Received for publication, March 8, 2007 , and in revised form, April 18, 2007.
* This work was supported by National Institutes of Health Grant PO1HL-20948 and the Perot Family Fund. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
The on-line version of this article (available at http://www.jbc.org) contains supplemental Tables S1 and S2 and Figs. S1 and S2.
1 Supported by a fellowship from the Canadian Institutes of Health Research.
2 To whom correspondence may be addressed: Dept. of Molecular Genetics, University of Texas Southwestern, 5323 Harry Hines Boulevard, Dallas, TX 75390-9046. Tel.: 214-648-6724; Fax: 214-648-7539; E-mail: Jonathan.cohen{at}utsouthwestern.edu.
3 To whom correspondence may be addressed: Dept. of Molecular Genetics, University of Texas Southwestern, 5323 Harry Hines Boulevard, Dallas, TX 75390-9046. Tel.: 214-648-6724; Fax: 214-648-7539; E-mail: Helen.hobbs{at}utsouthwestern.edu.
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