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232 ASSESSMENT OF IN VITRO PARAOXONASE 2 ENZYMATIC ACTIVITY: APPLICATIONS TO CARDIOVASCULAR DISEASE.
  1. V. Vanchinathan,
  2. N. Bourquard,
  3. S. T. Reddy
  1. Atherosclerosis Research Unit, Division of Cardiology, Department of Medicine, David Geffen School of Medicine at UCLA, Los Angeles, CA

Abstract

Oxidation of low-density lipoprotein (LDL) plays an integral role in the initiation and progression of atherosclerosis, an underlying cause for cardiovascular diseases. High-density lipoprotein (HDL) is antiatherogenic; HDL-associated proteins, including the paraoxonases (PON), contribute to the protective effects of HDL. The PON gene family consists of three members—PON 1, PON 2 and PON3—located on human chromosome 7q21.3-22.1. Biochemical studies have shown that all three PON proteins possess antioxidant properties against LDL in vitro. Although the physiological function and enzymatic substrates for the PON gene products remain unknown, the proteins possess a common enzyme activity: lactonase activity. In contrast to PON1 and PON3, PON2 is not associated with HDL in the circulation but instead remains intracellular. PON2 is ideal to study the mechanism of action of paraoxonases in vitro because it is associated with the plasma membrane fraction of cells. In the present study, we utilized cell culture models to determine whether the enzymatic activities of PON2 and two of its polymorphisms, A148M and C311S, correspond to their predicted roles in cardiovascular disease. Wild-type PON2, PON2 (A148M), and PON2 (C311S) were overexpressed in COS-7 cells using an adenoviral system. The activity of each enzyme was measured in a lactonase assay using 5-HETE lactone as the substrate. The results showed no significant difference in lactonase activity between the wild-type and variant enzymes. Based on these preliminary studies, we conclude that PON2 lactonase activity may not be associated with the predicted epidemiological link between PON2 polymorphisms and cardiovascular disease.

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