摘要
从玉米中克隆获得β-环化酶(lycopene β-cyclase,LCYb)和ε-环化酶(lycopene ε-cyclase,LCYe)基因,对编码产物进行生物学信息分析,通过体外大肠杆菌表达系统,借助于颜色互补及产物分析实验,探究了玉米LCYb和LCYe的催化功能特性。序列分析结果显示,玉米LCYb和LCYe的cDNA全长分别1 470 bp和1 611 bp,与高粱和小米物种的同源性均在90%以上。通过与谷胱甘肽巯基转移酶标签融合表达,成功纯化出LCYe和LCYb蛋白。颜色互补实验及高效液相产物分析结果表明,玉米LCYb具有β-环催化活性,可将番茄红素两端环化形成β-胡萝卜素,同时也具有微量的ε-环活性,可以通过中间体γ-胡萝卜素生成α-胡萝卜素;而玉米LCYe具有双ε-环活性可以同时催化番茄红素两端形成ε-胡萝卜素。本研究揭示了玉米番茄红素环化酶的催化特性,为探究玉米类胡萝卜素分子调控机制奠定了基础。
In this study,the lycopeneβ-cyclase(LCYb)and lycopeneε-cyclase(LCYe)genes were cloned from maize,and the encoded products were analyzed by bioinformatics methods.After expression in Escherichia coli,the catalytic properties of LCYb and LCYe from maize were explored by color complementation and product analysis experiments.The results of sequence analysis showed that the full-length cDNA of maize LCYb and LCYe were 1470 and 1611 bp,respectively,which were more than 90%homologous to those of sorghum and millet.LCYe and LCYb proteins were successfully purified by fusion expression with glutathione thiotransferase tags.The results of color complementation test and high performance liquid chromatography(HPLC)analysis showed that maize LCYb had catalytic activity onβ-ring,could cyclize both ends of lycopene to formβ-carotene,and had very weakε-ring catalytic activity,which could formα-carotene through the intermediateγ-carotene.Maize LCYe was also found to able to catalyze both ends of lycopene to formε-carotene.This study can lay a foundation for exploring the molecular mechanism of the regulation of maize carotenoid.
作者
王凡予
何伟伟
李大婧
郭庆启
罗浩
陆义珠
包怡红
张钟元
WANG Fanyu;HE Weiwei;LI Dajing;GUO Qingqi;LUO Hao;LU Yizhu;BAO Yihong;ZHANG Zhongyuan(College of Life Sciences,Northeast Forestry University,Harbin 150040,China;Institute of Agro-product Processing,Jiangsu Academy of Agricultural Sciences,Nanjing 210014,China;School of Food and Biological Engineering,Jiangsu University,Zhenjiang 212000,China)
出处
《食品科学》
EI
CAS
CSCD
北大核心
2023年第18期77-84,共8页
Food Science
基金
国家自然科学基金青年科学基金项目(31901710)。
关键词
玉米
番茄红素环化酶
基因克隆
颜色互补
功能验证
maize
lycopene cyclase
gene cloning
color complementation
functional verification
