‘A’-esterase activity in the lipoprotein fraction of sheep and human serum

Mackness, Michael I.; Hallam, S.D.; Walker, C.H.

doi:10.1042/bst0130135

Cited by 8 publications

(3 citation statements)

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“…The active fractions of M 2 containing PON1 [24] were discarded. The active fractions of M 1 (6)(7)(8)(9)(10)(11)(12)(13)(14)(15) were pooled, concentrated by ultrafiltration (Filtron) to a final volume of 10 ml and then dialysed overnight against buffer O. After use, the column was washed with 5 vol.…”

Section: Deae-sepharose Cl-6b/ion-exchange Chromatographymentioning

confidence: 99%

“…Paraoxonase (aryldialkylphosphatase, EC 3.1.8.1, PON1) is a calcium-dependent serum esterase that is synthesized by the liver and exhibits a broad-substrate specificity [1][2][3][4]. In serum, PON1 is closely associated with high-density lipoproteins [5,6] and recent studies have demonstrated the contribution of PON1 to the antioxidant protection conferred by high-density lipoproteins on low-density lipoprotein oxidation [4][5][6][7][8][9][10][11].…”

Section: Introductionmentioning

confidence: 99%

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Identification of paraoxonase 3 in rat liver microsomes: purification and biochemical properties

Rodrigo¹,

Gil²,

Hernández³

et al. 2003

Biochemical Journal

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Three paraoxonase genes (PON1, PON2 and PON3) have been described so far in mammals. Although considerable information is available regarding PON1, little is known about PON2 and PON3. PON3 has been isolated recently from rabbit serum [Draganov, Stetson, Watson, Billecke and La Du (2000) J. Biol. Chem. 275, 33435-33442] and liver [Ozols (1999) Biochem. J. 338, 265-275]. In the present study, we have identified the presence of PON3 in rat liver microsomes and a method for the purification to homogeneity is presented. PON3 has been purified 177-fold to apparent homogeneity with a final specific activity of 461 units/mg using a method consisting of seven steps: solubilization of the microsomal fraction, hydroxyapatite adsorption, chromatography on DEAE-Sepharose CL-6B, non-specific affinity chromatography on Cibacron Blue 3GA, two DEAE-cellulose steps and a final affinity chromatography on concanavalin A-Sepharose. SDS/PAGE of the final preparation indicated a single protein-staining band with an apparent molecular mass of 43 kDa. The isolated protein was identified by nanoelectrospray MS. Internal amino acid sequences of several peptides were determined and compared with those of human, rabbit and mouse PON3, showing a high similarity. Some biochemical properties of PON3 were also studied, including optimum pH, K(m) and heat and pH stability.

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Section: Deae-sepharose Cl-6b/ion-exchange Chromatographymentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Identification of paraoxonase 3 in rat liver microsomes: purification and biochemical properties

Rodrigo¹,

Gil²,

Hernández³

et al. 2003

Biochemical Journal

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“…In addition, the physiological role of A-esterases has yet not been clarified, since no natural substrates of these enzymes are known [12,13]. At present, paraoxon hydrolase (paraoxonase) has no known function in i o, although serum A-esterase has been reported to be associated with highdensity lipoproteins [14][15][16]. Therefore paraoxonase could be involved in atherogenesis, since this enzyme probably hydrolyses multiple oxygenated forms of polyunsaturated fatty acids at the sn-2 position of low-density-lipoprotein-associated phospholipids [17].…”

Section: Introductionmentioning

confidence: 99%

Purification and characterization of paraoxon hydrolase from rat liver

Rodrigo

Gil

Hernández

et al. 1997

Biochemical Journal

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Paraoxonase (paraoxon hydrolase), an enzyme that hydrolyses paraoxon (O,O-diethyl O-p-nitrophenyl phosphate), is located in mammals primarily in the serum and liver. Although considerable information is available regarding serum paraoxonase, little is known about the hepatic form of this enzyme. The present work represents the first study on the purification of rat liver paraoxonase. This enzyme has been purified 415-fold to apparent homogeneity with a final specific activity of 1370 units/mg using a protocol consisting of five steps: solubilization of the microsomal fraction, hydroxyapatite adsorption, chromatography on DEAE-Sepharose CL-6B, non-specific affinity chromatography on Cibacron Blue 3GA and anion exchange on Mono Q HR 5/5. The presence of Ca2+ and Triton X-100 in the buffers throughout the purification procedure was essential for maintaining enzyme activity. SDS/PAGE of the final preparation indicated a single protein-staining band with an apparent Mr of 45 000. N-terminal and internal amino acid sequences were determined and compared with those of paraoxonases from human and rabbit serum and mouse liver, showing a high similarity. The pH profile showed optimum activity at pH 8.5. The pH stability and heat inactivation of the enzyme were also studied. The Km for liver paraoxonase was 1.69 mM.

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Arylesterase

Schomburg¹,

Salzmann²

1991

Enzyme Handbook 3

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‘A’-esterase activity in the lipoprotein fraction of sheep and human serum

Cited by 8 publications

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Identification of paraoxonase 3 in rat liver microsomes: purification and biochemical properties

Identification of paraoxonase 3 in rat liver microsomes: purification and biochemical properties

Purification and characterization of paraoxon hydrolase from rat liver

Arylesterase

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