The development of human sera tests for HDL-bound serum paraoxonase PON1 and its lipo-lactonase activity

Leonid Gaidukov and Dan S. Tawfik

Department of Biochemistry, Weizmann Institute of Science, Rehovot 76100

Corresponding Author: leonid.gaydukov{at}weizmann.ac.il

Serum paraoxonase (PON1) is a lipo-lactonase that associates with 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 the total PON1 levels, irrespective of its 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 lipo-lactonase 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 aryl esterase tests weakly reflected the PON1-HDL levels (R=0.64), the lipo-lactonase test provided better correlation (R=0.80). These new tests indicate the levels and activity of PON1 in a physiologically relevant context, and the levels and quality of the HDL particles with which the enzyme is associated.

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