大蒜硫醚对葡萄糖-6-磷酸脱氢酶的抑制作用

Inhibitory Action of Garlic Thioether on Glucose 6 Phosphate Dehydrogenase

从大蒜中提取大蒜油,采用气相色谱-质谱联用仪(GC-MS)对其进行定性分析;采用酶活实时监测技术,揭示大蒜油、二烯丙基二硫醚(DADS)和二烯丙基三硫醚(DATS)对葡萄糖-6-磷酸脱氢酶(Glucose 6 Phosphate Dehydrogenase,G6PD)活性的抑制作用。酶动力学分析表明DADS/DATS对G6PD的抑制作用属于竞争性与非竞争性的混合性抑制。分子荧光技术分析表明DADS/DATS可对G6PD蛋白产生内源荧光淬灭,且为静态淬灭,提示DADS/DATS和G6PD蛋白结合位点数近似为1。热力学分析表明DADS-G6PD体系主要通过疏水作用结合,DATS-G6PD体系主要通过氢键和范德华力结合。利用3D和2D分子对接技术展示DADS/DATS可进入G6PD分子内部,且DADS与G6PD结合性能更好。DADS可与G6PD结合位点周围Ala-377、Val-376、Phe-216、Gly-222、Phe-221、Pro-223等疏水性氨基酸残基形成疏水作用,与Arg-219、Asn-218、Lys-275、Arg-215、Trp-225等极性氨基酸残基形成静电力,且以疏水作用为主。DATS可与G6PD结合位点周围Phe-253、Gly-254、Ala-335、Ile-472、Ile-255、Leu-469、Leu-305等疏水性氨基酸残基形成疏水相互作用,与Arg-175、Lys-476、Arg-257、Thr-334、Thr-333、Glu-473等极性氨基酸残基形成静电吸引。DATS中心位置的S可与Phe235形成氢键,键长为3.2?魡。疏水作用、静电力和氢键是DADS/DATS对G6PD的主要作用方式。

Garlic oil was extracted from garlic and qualitatively analyzed by Gas Chromatography-Mass Spectrometry(GC-MS).Using enzyme activity real-time monitoring technology,it was revealed that Garlic oil,diallyl disulfide(DADS)and diallyl trisulfide(DATS)had partial inhibitory effects on glucose 6 phosphate dehydrogenase(G6PD)activity,and the inhibition was a mixed inhibition of competitive and non-competitive by enzymatic reaction kinetics;Molecular fluorescence technique analysis further showed that DADS/DATS had static quenching effect on G6PD,and there was a binding site for DADS/DATS and G6PD protein;Thermodynamic analysis revealed that the DADS-G6PD system was mainly combined through hydrophobic interactions,and the DATS-G6PD system was mainly combined through hydrogen bond and van der Waals force;3D molecular docking and 2D molecular docking techniques further confirmed that DADS/DATS could enter the G6PD molecule,and the binding performance of DADS and G6PD was better;DADS could form hydrophobic interactions with Ala-377,Val-376,Phe-216,Gly-222,Phe-221,Pro-223 and other hydrophobic amino acid residues around the G6PD binding site,and interacted with Arg-219,Asn-218,Lys-275,Arg-215,Trp-225 and other polar amino acid residues formed electrostatic attraction,and hydrophobic interactions were dominant.DATS could form hydrophobic interactions with Phe-253,Gly-254,Ala-335,Ile-472,Ile-255,Leu-469,Leu-305 and other hydrophobic amino acid residues around the G6PD binding site,and interacted with Arg-175,Lys-476,Arg-257,Thr-334,Thr-333,Glu-473 and other polar amino acid residues formed electrostatic attraction.In addition,S in the center of DATS could form a hydrogen bond with Phe235 of G6PD,with a bond length of 3.2A.Hydrophobic interaction,electrostatic force and hydrogen bonding were the main modes of action of DADS/DATS on G6PD.