編碼S-腺苷-L-高半胱氨酸水解酶的草菇冷誘導基因Cor3功能驗證
Cold-Induced Volvariella volvacea Cor3 Gene Encodes an S-Adenosyl-L-homocysteine Hydrolase
【作者】 孫曉紅； 汪虹； 楊曉達； 馮愛萍； 陳明杰； 潘迎捷；
【Author】 SUN Xiaohong1,2,3,WANG Hong1,YANG Xiaoda4,FENG Aiping1,CHEN Mingjie1,PAN Yingjie3(1Institute of Edible Fungi,Shanghai Academy of Agricultural Sciences;Key Laboratory of Edible Fungi Resources and Utilization(South),Ministry of Agriculture,P.R.China;National Engineering Research Center of Edible Fungi;National R&D Center for Edible Fungi Processing;Key Laboratory of Agricultural Genetics and Breeding of Shanghai,Shanghai 201403,China;2College of Life Sciences,Nanjing Agricultural University,Nanjing,Jiangsu 210095,China,3Shanghai Ocean University,Shanghai 201306,China; 4Department of Chemical Biology,School of Pharmaceutical Science,Peking University,Beijing 100083)
【機構】 上海市農(nóng)業(yè)科學院食用菌研究所,農(nóng)業(yè)部南方食用菌資源利用重點實驗室,國家食用菌工程技術研究中心,國家食用菌加工技術研發(fā)分中心,上海市農(nóng)業(yè)遺傳育種重點開放實驗室； 南京農(nóng)業(yè)大學生命技術學院； 上海海洋大學； 北京大學醫(yī)學部藥學院；
【摘要】 將草菇(Volvariella volvacea)冷誘導基因Cor3cDNA序列經(jīng)EcoR I酶切克隆到原核表達載體pPROK-1中,轉化大腸桿菌(Escherichia coli)JM109,經(jīng)異丙基-β-D-硫代吡喃半乳糖苷(IPTG)誘導表達、分離純化發(fā)現(xiàn)該蛋白的分子量約為43kD,等電聚焦電泳結果表明,該蛋白的等電點為6.1,HPLC檢測說明合成方向的酶活性為每毫克蛋白每分鐘4.5μmol,DNTB與同型半胱氨酸顏色反應測定水解方向的酶活性為每毫克蛋白每分鐘0.7μmol,根據(jù)上述特性可以確定草菇冷誘導基因Cor3所編碼的蛋白是S-腺苷同型半胱氨酸水解酶(SAHH)。
【Abstract】 The full-length cDNA sequence of Volvariella volvacea Cor3 gene was digested with EcoRI restriction enzyme,ligated to the expression vector pPROK-1,and the recombinant plasmid was transformed into Escherichia coli JM109.The encoded protein was over-expressed in the transformed bacteria by addition of 0.5 mmol/L IPTG to the culture medium and cell-free extracts were prepared by sonication.DEAE-cellulose ion-exchange,Sephacryl S-200 gel filtration and QAE-Sepharose negative ion exchange chromatogr...
【關鍵詞】 S-腺苷同型半胱氨酸水解酶基因； 草菇； 冷誘導基因；
【Key words】 S-adenosyl-L-homocysteine hydrolase gene； Volvariella volvacea； cold induced gene；
【基金】 草菇基因組和功能基因組數(shù)據(jù)庫共享平臺建設(編號:10dz2212400);上海市科學技術委員會重點項目的部分研究內(nèi)容
【分類號】S646.13