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Originally published In Press as doi:10.1194/jlr.D600045-JLR200 on April 13, 2007
Papers In Press, published online ahead of print July 1, 2007
J. Lipid Res., doi:10.1194/jlr.D600045-JLR200
Journal of Lipid Research, Vol. 48, 1637-1646, July 2007
Copyright © 2007 by American Society for Biochemistry and Molecular Biology
The development of human sera tests for HDL-bound serum PON1 and its lipolactonase activity
Leonid Gaidukov and
Dan S. Tawfik1
Department of Biological Chemistry, Weizmann Institute of Science, Rehovot 76100, Israel
The online version of this article (available at http://www.jlr.org) contains supplementary data in the form of one table and five figures.
Published, JLR Papers in Press, April 13, 2007.
1 To whom correspondence should be addressed. e-mail: tawfik{at}weizmann.ac.il
Serum paraoxonase (PON1) is a lipolactonase that associates with HDL-apolipoprotein A-I (HDL-apoA-I) and thereby plays a role in the prevention of atherosclerosis. Current sera tests make use of promiscuous substrates and provide no indications regarding HDL-PON1 complex formation. We developed new enzymatic tests that detect total PON1 levels, irrespective of HDL status and R/Q polymorphism, as well as the degree of catalytic stimulation and increased stability that follow PON1's tight binding to HDL-apoA-I. The tests are based on measuring total PON1 levels with a fluorogenic phosphotriester, measuring the lipolactonase activity with a chromogenic lactone, and assaying the enzyme's chelator-mediated inactivation rate. The latter two are affected by tight HDL binding and thereby derive the levels of the serum PON1-HDL complex. We demonstrate these new tests with a group of healthy individuals (n = 54) and show that the levels of PON1-HDL vary by a factor of 12. Whereas the traditionally applied paraoxonase and arylesterase tests weakly reflect PON1-HDL levels (R = 0.64), the lipolactonase test provides better correlation (R = 0.80). These new tests indicate the levels and activity of PON1 in a physiologically relevant context as well as the levels and quality of the HDL particles with which the enzyme is associated.
Supplementary key words blood tests lactones lipoproteins high density lipoprotein-bound enzymes enzyme stability atherosclerosis paraoxonase Abbreviations: apoA-I, apolipoprotein A-I; DEPCyMC, 7-O-diethyl phosphoryl 3-cyano 4-methyl 7-hydroxycoumarin; DTNB, 5,5'-dithio-bis-2-nitrobenzoic acid; PON1, serum paraoxonase; TBBL, 5-thiobutyl butyrolactone

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Copyright © 2007 by the American Society for Biochemistry and Molecular Biology.
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