Molecular basis for LDL receptor recognition by PCSK9

Abstract

hypercholesterolemia ͉ proprotein convertase ͉ crystal structure P roprotein convertase (PC) subtilisin/kexin type 9 (PCSK9) is a serine protease of the PC family that has profound effects on plasma low-density lipoprotein (LDL)-cholesterol (LDL-C) levels through its ability to mediate LDL receptor (LDLR) protein degradation (1, 2). The link between PCSK9 and plasma LDL-C levels was first established by the discovery of missense mutations in PCSK9 that were present in patients with an autosomal dominant form of familial hypercholesterolemia (FH) (3). These mutations were speculated to result in a gain-offunction of PCSK9 owing to their mode of inheritance. Subsequently, loss-of-function mutations in PCSK9 were discovered that are associated with low plasma LDL-C levels and significantly reduced coronary heart disease (CHD) (4, 5).The overall domain structure of PCSK9 is similar to other PC family members. It includes a signal peptide, followed by a prodomain, a subtilisin-like catalytic domain, and a variable C-terminal domain (6). The prodomain serves a dual role as a chaperone for folding and as an inhibitor of catalytic activity (7-9). Autocatalysis between Gln-152 and Ser-153 (VFAQ:SIP) separates the prodomain from the catalytic domain, but the prodomain remains bound, occluding the catalytic site (10, 11). For other PC family members, a second catalytic cleavage is required to release the prodomain, which unmasks the catalytic site, resulting in an active protease (7). No site of secondary cleavage has been identified that activates PCSK9.The crystal structure of apo-PCSK9 revealed a tightly bound prodomain that is predicted to render the active site inaccessible to exogenous substrates (12-14). The structure of the PCSK9 prodomain and catalytic domain is similar to that of other subtilisin-like serine proteases. The C-terminal domain of PCSK9 contains three six-stranded ␤-sheet subdomains arranged with quasi-threefold symmetry. This domain shares structural homology to the adipokine resistin and has been speculated to mediate protein-protein interactions (12-14).Recent studies have provided insights into the site and mode of PCSK9's action. Addition of recombinant PCSK9 to the medium of cultured hepatocytes results in a dramatic reduction in LDLR number. PCSK9 binds directly to the LDLR on the cell surface and PCSK9-stimulated degradation of the LDLR requires autosomal recessive hypercholesterolemia (ARH), an adaptor protein needed for internalization of LDLRs (11). The affinity of PCSK9 binding to the LDLR is enhanced at acidic pH, suggesting that PCSK9 binds more avidly to LDLRs in the lysosomal/endosomal compartments (12,15,16). One gain-offunction mutant, Asp-374-Tyr, was Ϸ10-fold more active than wild-type PCSK9 in mediating degradation of LDLRs, owing to an Ϸ5-to 30-fold increased affinity of PCSK9 for the LDLR (11,12,15). Although autocatalysis of PCSK9 is required for proper folding and secretion, catalytic activity was not required for PCSK9-mediated LDLR degradation when added to cultured cells (...

Mutations and polymorphisms in the proprotein convertase subtilisin kexin 9 (PCSK9) gene in cholesterol metabolism and disease

“…The binding site for the LDLR EGF-A domain resides on the surface of PCSK9 that is formed primarily by residues 367 to 381. Key interactions with EGF-A are made by Arg-194 and Asp-238 of PCSK9 [Kwon et al, 2008]. Moreover, PCSK9 contains a fully-folded C-terminal cysteine-rich domain showing a distinct structural similarity to the resistin homotrimer, a small cytokine associated with obesity and diabetes .…”

“…Finally, the low plasma LDL-C levels and the reduction of CHD displayed in PCSK9 loss-of-function mutation carriers, which seem furthermore healthy, indicate that the inhibition of PCSK9 should be an effective target in the treatment of hypercholesterolemia with no known adverse side effects. The recent elucidation of the crystal structure of PCSK9 [Cunningham et al, 2007;Piper et al, 2007], especially in complex with the LDL receptor [Kwon et al, 2008], should provide essential clues for the identification of small pharmacological molecules or antibodies that might block the interaction of PCSK9 and the LDLR and thus inhibit PCSK9 function and constitute new cholesterol-lowering drugs [Kwon et al, 2008;Grefhorst et al, 2008].…”

Evaluation of reported pathogenic variants and their frequencies in a Japanese population based on a whole-genome reference panel of 2049 individuals

“…The protein is synthesized as a soluble zymogen that undergoes autocatalytic cleavage in the endoplasmic reticulum. The mature protein binds to the EGF-A domain of lipoprotein receptors [70,71], abolishes their functions, and raises the level of cholesterol in the blood stream. Furthermore, some gain-offunction mutations increase the binding affinity of PCSK9 and lipoprotein receptors, thus resulting in the degradation of LDLR, inefficient incorporation of cholesterol in the liver cells, and higher cholesterol levels in the blood stream.…”

Atomic structure of the autosomal recessive hypercholesterolemia phosphotyrosine-binding domain in complex with the LDL-receptor tail

“…Moreover, so far only the ectodomain of LDLR has received structural attention (18)(19)(20)(21)(22)(23)(24), and no structural data for the LDLR tail are available to establish the molecular basis for its interaction with ARH and for hypercholesterolemia resulting from internalization defects.…”