文献类型：期刊文献

中文题名：产酸克雷伯菌中水解酶的表达及其在果蔬表面多菌灵去除中的应用

英文题名：Expression of a Novel Hydrolase from Klebsiella oxytoca for Carbendazim Residue Degradation on the Surface of Fruit and Vegetable

作者：陈慧勤[1];崔婷[2]

第一作者：陈慧勤

机构：[1]苏州大学附属第一医院临床教学办,江苏苏州215006;[2]北京联合大学图书馆,北京100101

第一机构：苏州大学附属第一医院临床教学办,江苏苏州215006

年份：2019

卷号：40

期号：1

起止页码：116-120

中文期刊名：食品工业科技

外文期刊名：Science and Technology of Food Industry

收录：CSTPCD;;北大核心:【北大核心2017】；

语种：中文

中文关键词：多菌灵;水解酶;产酸克雷伯菌;农残降解

外文关键词：carbendazim;hydrolase;Klebsiella oxytoca;pesticide residue degradation

摘要：为研究酶法清除果蔬表面多菌灵残留效果,从多菌灵污染的土壤中筛选多菌灵降解细菌,并从中克隆多菌灵水解酶,表达纯化后制备为酶制剂,催化果蔬表面多菌灵降解。实验结果表明:通过筛选,从土壤中得到了一株可利用多菌灵为唯一碳源的产酸克雷伯菌Klebsiella oxytoca KO-1。以基因组为模板使用简并引物PCR扩增,得到了一段837 bp的水解酶编码基因。将该基因连接至pET-28a并转化至大肠杆菌BL21中,重组菌成功表达出了28 kDa的水解酶KY-1。使用镍柱蛋白纯化法,成功实现了KY-1的纯化并制备了KY-1酶水剂。经液质检测,KY-1将多菌灵催化为2-氨基苯并咪唑,其纯化蛋白酶活达到36.8 U/mg。将KY-1酶水剂喷洒至果蔬表面后,成功提高了黄瓜、草莓、西红柿和土豆表面多菌灵残留的降解率,最高可达37.3%。本研究成功筛选了一株多菌灵降解菌,并制备了可用于果蔬表面多菌灵降解的酶制剂,为酶法清除农药残留奠定了基础。
In order to explore enzymatic method for pesticide residue degradation on surface of fruits,a novel carbendazim degradation bacteria was isolated and the carbendazim hydrolase coding gene was amplified.The results showed that a novel Klebsiella oxytoca KO-1 which was able to survive with carbendazim as the sole carbon source was screened and identified. Moreover,a 837bp hydrolase coding gene was amplified by PCR using degenerate primers,and hydrolase KY-1 that catalyzing carbendazim degradation was expressed and purified in E.coli BL 21.KY-1successfully catalyzed carbendazim into 2-aminobenzimidazole depends on HPLC-MS analysis results,and the relative enzymatic activity was 36.8U/mg.The enzyme preparation of KY-1 successfully accelerated carbendazim degradation rate on the surface of cucumber,potato,tomato and strawberry,and the highest acceleration rate was 37.3%.In conclusion,a carbendazim degradation bacterium was isolated and the enzyme preparation was prepared.Moreover,this work lay the foundation for the enzymatic restoration of pesticide contaminated fruits.