Atatürk University, Science Faculty, Department of Chemistry, Biochemistry Division, Erzurum, Turkey.
Chem Biol Drug Des. 2010 Dec;76(6):552-8. doi: 10.1111/j.1747-0285.2010.01036.x. Epub 2010 Oct 11.
Paraoxonase 1 (PON1), a high-density lipoprotein (HDL)-associated esterase, is known to mediate antioxidant and antiatherogenic properties. Purification of PON1 has been challenging for a long time. Here, we report a novel purification technique for this enzyme, which allowed us to obtain human serum paraoxonase 1 (hPON1) using straightforward chromatographic methods, such as Diethylaminoethyl-Sephadex anion exchange chromatography and Sepharose 4B-4-phenylazo-2-naphthaleneamine hydrophobic interaction chromatography. We purified the enzyme 302-fold with a final specific activity of 4775 U/mg and a yield of 32%. Furthermore, we examined the in vitro effects of some sulfonamide derivatives, such as sulfacetamide, homosulfanilamide (mafenide), sulfosalazine, furosemide, acetazolamide, and 1,3,4-thiadiazole-2-sulfonamide on the enzyme activity to better understand the inhibitory properties of the molecules. The six sulfonamides dose-dependently decreased the activity of hPON1 with inhibition constants in the millimolar-micromolar range. This study provides an efficient method, which may be useful for other enzymes such as those related to acetylcholinesterase. It also demonstrates the off-target activity of sulfonamides.
对氧磷酶 1（PON1）是一种高密度脂蛋白（HDL）相关的酯酶，已知具有抗氧化和抗动脉粥样硬化的特性。很长一段时间以来，对 PON1 的纯化一直具有挑战性。在这里，我们报告了一种用于该酶的新型纯化技术，该技术使我们能够使用简单的色谱方法（如二乙基氨基乙基-葡聚糖阴离子交换色谱法和琼脂糖 4B-4-对氨基偶氮-2-萘酚胺疏水相互作用色谱法）获得人血清对氧磷酶 1（hPON1）。我们将酶纯化了 302 倍，最终比活性为 4775 U/mg，收率为 32%。此外，我们还研究了一些磺胺衍生物，如磺胺醋酰、甲芬那酸（氨苯磺胺）、柳氮磺胺吡啶、呋塞米、乙酰唑胺和 1,3,4-噻二唑-2-磺胺对酶活性的体外影响，以更好地了解分子的抑制特性。六种磺胺衍生物剂量依赖性地降低了 hPON1 的活性，抑制常数在毫摩尔-微摩尔范围内。本研究提供了一种有效的方法，可能对其他酶（如与乙酰胆碱酯酶相关的酶）有用。它还证明了磺胺类药物的非靶标活性。
