Isolation and characterization of the circulating truncated form of PCSK9.
Proprotein convertase subtilisin-kexin type 9 (PCSK9) is a secreted protein which regulates serum LDL cholesterol. It circulates in human and rodent serum in an intact form and a major truncated form. Previous in vitro studies involving the expression of human PCSK9 genetic variants and in vivo studies of furin knockout mice suggest that the truncated form is a furin cleavage product. However, the circulating truncated form of PCSK9 has not been isolated and characterized. Utilizing antibodies which bind to either the catalytic domain or the C-terminal domain of PCSK9, the truncated PCSK9 was isolated from serum. MS was used to determine that this form of PCSK9 is a product of in vivo cleavage at Arg218 resulting in pyroglutamic acid formation of the nascent N terminus corresponding to Gln219 of intact PCSK9. We also determined that the truncated PCSK9 in serum lacked the N-terminal segment which contains amino acids critical for LDL receptor binding. A truncated PCSK9, expressed and purified from HEK293 cells with identical composition as the circulating truncated protein, was not active in inhibition of LDL uptake by HepG2 cells. These studies provide a definitive characterization of the composition and activity of the truncated form of PCSK9 found in human serum.
Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, IN 46285.
Bomie
B
Han
Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, IN 46285.
Patrick I
PI
Eacho
Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, IN 46285.
Michael D
MD
Knierman
Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, IN 46285.
Jason S
JS
Troutt
Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, IN 46285.
Robert J
RJ
Konrad
Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, IN 46285.
Xiaohong
X
Yu
Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, IN 46285.
Krista M
KM
Schroeder
D000818:Animals; D057809:HEK293 Cells; D056945:Hep G2 Cells; D006801:Humans; D051379:Mice; D018345:Mice, Knockout; D000071449:Proprotein Convertase 9 / Q000097:blood / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D000072417:Protein Domains
C472125:PCSK9 protein, human; D000071449:Proprotein Convertase 9
furin; low density lipoprotein cholesterol; low density lipoprotein receptor; monoclonal antibodies; proprotein convertase subtilisin-kexin type 9; proteolytic cleavage