酯酶D的表达纯化和激活剂筛选  

A study on expression,purification and activator screening of Esterase D

作　　者：岑雪艳 宋嫚 李梓灵 贾晓悦 张柔 张霓 陈新鹏 CEN Xue-yan;SONG Man;LI Zi-ling;JIA Xiao-yue;ZHANG Rou;ZHANG Ni;CHEN Xin-peng(College of Life Science,Hubei Normal University,Huangshi 435002,China)

机构地区：[1]湖北师范大学生命科学学院,湖北黄石435002

出　　处：《湖北师范大学学报（自然科学版）》2023年第2期15-19,共5页Journal of Hubei Normal University：Natural Science

摘　　要：酯酶D(Esterase D,ESD)在解毒过程中起重要作用,在临床上经常是癌症和遗传疾病的生物标志物,在多种疾病中其活性降低。提高ESD的活性是治疗相关疾病的一种有效策略,但是目前还没有该蛋白高效的小分子激活剂。将构建好的ESD表达质粒转化到BL21(DE3)感受态细胞中并诱导表达,通过镍离子亲和层析方法得到高活性的ESD.通过检测多种小分子对ESD活性的影响,筛选出对ESD具有激活作用的一种小分子—Dolutegravir.分子对接计算结果显示,Dolutegravir与人ESD的最低结合自由能为-8.9 kcal/mol,结合的活性位点为Asn-133、Pro-140、Tyr-166和Lys-162残基。筛选ESD的小分子激活剂,不仅有助于阐明ESD的激活机理,还可以为后面的药物开发作参考。Esterase D(ESD)plays an important role in detoxification and is often used as a biomarker for many types of cancer and genetic diseases.Since its activity can be reduced in many diseases,increasing its activity is an effective strategy for the treatment of related diseases.However,no protein activation molecules have been found in clinical trials.This study transformed ESD expression plasmid into BL21(DE3)competent cells to induce expression,and obtainedpurified ESD with high activity with the application of nickel ion affinity chromatography.Then,it screened out Dolutegravir,a small activation molecule by detecting the effects of different small molecules on ESD activity.Results gained by the molecular docking approach indicated the minimum binding free energy between Dolutegravir and ESD was-8.9kcal/mol andthe binding active sites were Asn-133,Pro-140,Tyr-166,and Lys-162 residues.The screening of ESD activation molecules can not only help to clarify the activation mechanism of ESD,but also provide references for the development of drugs in the future.

关 键 词：ESD 蛋白质表达 蛋白质纯化 分子对接 激活剂 

分 类 号：Q5[生物学—生物化学]