The effect of some detergents and stabilizing agents on plasma membrane phosphatidate phosphohydrolase of rat liver

Esfandiar Heidarian; Bahram Haghighi; Esmat Jafari Dehkordy

Journal of Kermanshah University of Medical Sciences

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The effect of some detergents and stabilizing agents on plasma membrane phosphatidate phosphohydrolase of rat liver

Author(s):

Esfandiar Heidarian^1,*,

Bahram Haghighi²,

Esmat Jafari Dehkordy³

1Dept. of Biochemistry, School of Medicine, Ilam University of Medical Sciences, Ilam, Iran

2Dept. of Biochemistry, School of Medicine, Esfahan University of Medical Sciences, Esfahan, Iran

3Share-Kord Health Center, Share-Kord University of Medical Sciences, Shahre-kord, Iran

Journal of Kermanshah University of Medical Sciences:Vol. 13, issue 1; e79824

Published online:Jun 19, 2009

Article type:Research Article

Received:Jun 09, 2008

Accepted:Jun 02, 2009

How to Cite:Esfandiar HeidarianBahram HaghighiEsmat Jafari DehkordyThe effect of some detergents and stabilizing agents on plasma membrane phosphatidate phosphohydrolase of rat liver.J Kermanshah Univ Med Sci.13(1):e79824.

Abstract

Background: Phosphatidate phosphohydrolase (PAP) catalyzes the dephosphorylation of phosphatidic acid to yield Pi and diacylglycerol. Two different forms of PAP have been reported in rat hepatocyte: PAP1 that participates in the synthesis of phospholipids and triacylglycerols and PAP2 which is involved in lipid signaling pathways. Two isoforms of PAP2 are PAP2a and PAP2b. This study examines the effect of detergents such as Tween 80 lubrol, PX and CTAB as well as stabilizing factors including trehalose, sucrose, and albumin on the stability and activity of PAP2b.

Methods: 14 Wistar rats weighing between 200-250 grams were used in the study. PAP2b was purified from liver plasma membrane by solubilizing with n-octyle glucoside and through several chromatography stages. Gel electrophoresis (SDSPAGE) was performed in 10%gel slab in order to determine the purity level and to measure the molecular weight and number of the enzyme subunit. The effect of trehalose, sucrose, and albumin was examined on the stimulation of enzyme activity in different concentrations.

Results: The specific activity of purified enzyme was 7350 mU/mg protein. The purified enzyme showed a single band on SDS-PAGE with a MW of about 33.8 kDa. The enzyme was approximately activated 3times by lubrol PX and Tween 80 both at 3 mM. The activation by CTAB occurred at 1Mm.Trehalose, sucrose and albumin had the most stability effect on PAP2b in concentration of 3, 7 and 10 percent respectively.

Conclusions: Tween 80, lubrol PX, and CTAB have the ability to activate PAP2b. In case a nondetergent agent is required to stabalize PAP2b trehalose is preferred to sucrose and albumin. The lubrol PX has a higher potential stimulatory effect to activate PAP2b. compared to other ionic and nonionic detergents.

Keywords

phosphatidate phosphohydrolase

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