Department of Biochemistry, Faculty of Life Sciences, Aligarh Muslim University, Aligarh, 202002, India.
Department of Biochemistry, Faculty of Life Sciences, Aligarh Muslim University, Aligarh, 202002, India.
Environ Toxicol Pharmacol. 2018 Jul;61:52-60. doi: 10.1016/j.etap.2018.05.019. Epub 2018 May 24.
Pesticides are being used globally to improve agricultural production. They are applied specifically to combat with pathogens that are a major threat for reduced optimum yield of crops. This study was carried out to see the effect of commercially used pesticides on a specific plant protein viz. phytocystatin isolated from yellow mustard seeds (YMP). Phytocystatin is a thiol proteinase inhibitor, which regulates endogenous and exogenous cysteine proteinases and plays a vital physiological role in plants. Different classes of pesticides like fungicide (iprodione) of dicarboximide class and an insecticide (malathion) of class organophosphate are retorted for our study. In the presence of these pesticides, biophysical and biochemical changes were observed in phytocystatin. These changes were evaluated making use of caseinolytic activity assay, UV-vis spectroscopy, fluorescence spectroscopy, FTIR, and circular dichroism. Isothermal titration calorimetry was employed to see interaction pattern of these pesticides with phytocystatin. The results obtained clearly depict that the pesticides bind with the phytocystatin thereby changing its native conformation and reducing its intrinsic property of inhibition on cysteine proteinase as evident by reduced anti-papain inhibition in the presence of pesticides. Furthermore, CD and FTIR spectroscopy results clearly show a decrease in α-helical content upon interaction with malathion and iprodione. Among the two pesticides, iprodione has far more pronounced effect on YMP evident from striking changes in UV, Fluorescence, CD and FTIR spectroscopy. 2,4-dinitrophenylhydrazine spectrophotometric assay was also carried out to check production of ROS, generation of ROS was observed in the presence of these pesticides thus implying that ROS might be responsible for changes in native structure of phytocystatin induced by pesticides.
农药被全球广泛用于提高农业产量。它们专门用于对抗病原体，这些病原体是降低作物最佳产量的主要威胁。这项研究旨在观察商业上使用的农药对一种特定植物蛋白（即从黄芥菜种子中分离出的植源性胱抑素）的影响。植源性胱抑素是一种巯基蛋白酶抑制剂，可调节内源性和外源性半胱氨酸蛋白酶，并在植物中发挥重要的生理作用。我们研究了不同类别的农药，如二羧酰亚胺类杀菌剂（异菌脲）和有机磷类杀虫剂（马拉硫磷）。在这些农药的存在下，观察到植源性胱抑素的生物物理和生化变化。利用酪蛋白水解活性测定、紫外-可见光谱、荧光光谱、傅里叶变换红外光谱和圆二色性评估这些变化。采用等温滴定量热法观察这些农药与植源性胱抑素的相互作用模式。结果清楚地表明，这些农药与植源性胱抑素结合，从而改变其天然构象，并降低其对半胱氨酸蛋白酶的内在抑制特性，这从存在农药时对半 Papain 的抑制减少得到证明。此外，CD 和 FTIR 光谱结果清楚地表明，与马拉硫磷和异菌脲相互作用后α-螺旋含量减少。在这两种农药中，异菌脲对 YMP 的影响更为明显，从对 UV、荧光、CD 和 FTIR 光谱的显著变化中可以明显看出。还进行了 2,4-二硝基苯肼分光光度法测定以检查 ROS 的产生，在这些农药的存在下观察到 ROS 的产生，这意味着 ROS 可能是由农药引起的植源性胱抑素天然结构变化的原因。
