Tandem purification of two HDL-associated partner proteins in human plasma, paraoxonase (PON1) and phosphate binding protein (HPBP) using hydroxyapatite chromatography

Renault, Frédérique; Chabrière, Éric; Andrieu, Jean-Pierre; Dublet, Bernard; Masson, Patrick; Rochu, Daniel

doi:10.1016/j.jchromb.2006.03.029

Cited by 97 publications

(72 citation statements)

References 26 publications

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“…Human paraoxonase (PON1) purified to homogeneity [13] was tested for AAA activity with o-NTFNAC as described above. Because the concentration of PON1 in human plasma is about 1000 fold less than that of albumin, activity assays with PON1 were carried out with an enzyme concentration less than 0.1 mM.…”

Section: Measurement Of Aaa Activity Of Human Paraoxonasementioning

confidence: 99%

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Aryl acylamidase activity of human serum albumin witho-nitrotrifluoroacetanilide as the substrate

Masson

Froment

Darvesh

et al. 2007

Journal of Enzyme Inhibition and Medicinal Chemistry

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Albumin is generally regarded as an inert protein with no enzyme activity. However, albumin has esterase activity as well as aryl acylamidase activity. A new acetanilide substrate, o-nitrotrifluoroacetanilide (o-NTFNAC), which is more reactive than the classical o-nitroacetanilide, made it possible to determine the catalytic parameters for hydrolysis by fatty-acid free human serum albumin. Owing to the low enzymatic activity of albumin, kinetic studies were performed at high albumin concentration (0.075 mM). The albumin behavior with this substrate was Michaelis-Menten like. Kinetic analysis was performed according to the formalism used for catalysis at high enzyme concentration. This approach provided values for the turnover and dissociation constant of the albumin-substrate complex: k cat ¼ 0.13^0.02 min2 1 and K s ¼ 0.67^0.04 mM. MALDI-TOF experiments showed that unlike the ester substrate p-nitrophenyl acetate, o-NTFNAC does not form a stable adduct (acetylated enzyme). Kinetic analysis and MALDI-TOF experiments demonstrated that hydrolysis of o-NTFNAC by albumin is fully rate-limited by the acylation step (k cat ¼ k 2 ). Though the aryl acylamidase activity of albumin is low (k cat /K s ¼ 195 M 21 min 21 ), because of its high concentration in human plasma (0.6 -1 mM), albumin may participate in hydrolysis of aryl acylamides through second-order kinetics. This suggests that albumin may have a role in the metabolism of endogenous and exogenous aromatic amides, including drugs and xenobiotics.

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Section: Measurement Of Aaa Activity Of Human Paraoxonasementioning

confidence: 99%

“…3.5.1. 13) activity of albumin was demonstrated with o-nitroacetanilide as the substrate [7]. The organophosphate diisopropylfluorophosphate progressively inhibited the AAA activity of albumin.…”

Section: Introductionmentioning

confidence: 99%

Aryl acylamidase activity of human serum albumin witho-nitrotrifluoroacetanilide as the substrate

Masson

Froment

Darvesh

et al. 2007

Journal of Enzyme Inhibition and Medicinal Chemistry

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“…Activity measurement is based on establishing the changes in absorption of formed p-nitrophenol as a result of the reaction of PON1 with paraoxon at 412 nm. p-Nitrophenol the molar extinction coefficient is e = 18.290M −1 cm −1 and an enzyme unit is number of micromole of hydrolysis paraoxon in per minute [41][42][43].…”

Section: 22mentioning

confidence: 99%

Synthesis and evaluation of novel 2-[(1,2,4-triazol-3-yl)thio]acetamide derivatives as potential serum paraoxonase-1 (PON1) activators

Yurttaş¹,

Küçükoğlu²,

Nadaroğlu³

et al. 2017

mpj

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Coronary artery disease and low-density lipoprotein (LDL) levels in the blood have long been known to be associated with peripheral vascular diseases. Paraoxonase-1 (PON1) enzyme is related to serum levels of high-density lipoprotein (HDL) and protects LDL from oxidation which may result in development of microvascular disease in diabetes. The enzyme has a major role in the prevention of atherosclerosis besides antioxidant properties. Additionally, PON1 is important in the detoxification of organophosphate insecticides from the body. In this study, we aimed to synthesize highly active new compounds which can be a drug candidate and evaluate their effects on PON1 activity. Nine novel triazole compounds bearing thioacetamide moiety (5a-i) were synthesized and their in vitro PON1 activity was investigated. The PON1 enzyme was purified from human serum using ammonium sulfate precipitation method. Also, it was further purified using Sepharose 4B-L-tyrosine-1-naphthylamine affinity chromatography. Among the synthesized triazole compounds, 5b, 5c, 5f and 5h have been determined to increase PON1 activity, remarkably. Compounds 5b, 5c, 5f and 5h bearing 5-nitrothiazole, benzothiazole, 6-ethoxybenzothiazole and 6-florobenzothiazole moieties could be considered to proceed in vivo investigations which is a further stage for a drug candidate.

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“…It is e = 18,290 M −1 cm −1 at pH 10.5. 13 One enzyme unit was defined as the amount of enzyme that catalyzes the hydrolysis of 1 μmol of substrate at 25°C.…”

Section: Pon Activity Assaymentioning

confidence: 99%

Impacts of some antibiotics on human serum paraoxonase 1 activity

Dilek

Küfrevioğlu

Beydemır

2012

Journal of Enzyme Inhibition and Medicinal Chemistry

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Tandem purification of two HDL-associated partner proteins in human plasma, paraoxonase (PON1) and phosphate binding protein (HPBP) using hydroxyapatite chromatography

Cited by 97 publications

References 26 publications

Aryl acylamidase activity of human serum albumin witho-nitrotrifluoroacetanilide as the substrate

Aryl acylamidase activity of human serum albumin witho-nitrotrifluoroacetanilide as the substrate

Synthesis and evaluation of novel 2-[(1,2,4-triazol-3-yl)thio]acetamide derivatives as potential serum paraoxonase-1 (PON1) activators

Impacts of some antibiotics on human serum paraoxonase 1 activity

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