Identification of the active form of endothelial lipase, a homodimer in a head-to-tail conformation.
Endothelial lipase (EL) is a member of a subfamily of lipases that act on triglycerides and phospholipids in plasma lipoproteins, which also includes lipoprotein lipase and hepatic lipase. EL has a tropism for high density lipoprotein, and its level of phospholipase activity is similar to its level of triglyceride lipase activity. Inhibition or loss-of-function of EL in mice results in an increase in high density lipoprotein cholesterol, making it a potential therapeutic target. Although hepatic lipase and lipoprotein lipase have been shown to function as homodimers, the active form of EL is not known. In these studies, the size and conformation of the active form of EL were determined. Immunoprecipitation experiments suggested oligomerization. Ultracentrifugation experiments showed that the active form of EL had a molecular weight higher than the molecular weight of a simple monomer but less than a dimer. A construct encoding a covalent head-to-tail homodimer of EL (EL-EL) was expressed and had similar lipolytic activity to EL. The functional molecular weights determined by radiation inactivation were similar for EL and the covalent homodimer EL-EL. We previously showed that EL could be cleaved by proprotein convertases, such as PC5, resulting in loss of activity. In cells overexpressing PC5, the covalent homodimeric EL-EL appeared to be more stable, with reduced cleavage and conserved lipolytic activity. A comparative model obtained using other lipase structures suggests a structure for the head-to-tail EL homodimer that is consistent with the experimental findings. These data confirm the hypothesis that EL is active as a homodimer in head-to-tail conformation.
Griffon
Nathalie
N
Institute for Translational Medicine and Therapeutics, Cardiovascular Institute, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania 19104, USA.
Jin
Weijin
W
Petty
Thomas J
TJ
Millar
John
J
Badellino
Karen O
KO
Saven
Jeffery G
JG
Marchadier
Dawn H
DH
Kempner
Ellis S
ES
Billheimer
Jeffrey
J
Glick
Jane M
JM
Rader
Daniel J
DJ
D002460:Cell Line; D019281:Dimerization; D006801:Humans; D008049:Lipase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008968:Molecular Conformation; D011487:Protein Conformation; D015202:Protein Engineering
C410382:LIPG protein, human; D008049:Lipase