犹他游动放线菌SW1311中阿库来菌素A酰基酶在棘白菌素类抗真菌药物合成中的应用(英文).  

Application of an aculeacin A acylase from Actinoplanes utahensis SW1311 in syntheses of echinocandins

作　　者：冯国栋 许永锋 陈建烽 赵万忠 王欣 张小晟 江辉[3] FENG Guodong;XU Yongfeng;CHEN Jianfeng;ZHAO Wanzhong;WANG Xin;ZHANG Xiaosheng;JIANG Hui(Hangzhou Huadong Medicine Group Pharmaceutical Research Institute Co., Ltd., Hangzhou 310011, China;Qiushi College, Zhejiang University, Hangzhou 310058, China;College of Life Sciences, Zhejiang University, Hangzhou 310058, China)

机构地区：[1]华东医药集团新药研究院有限公司,杭州310011 [2]浙江大学求是学院,杭州310058 [3]浙江大学生命科学学院,杭州310058

出　　处：《浙江大学学报（农业与生命科学版）》2017年第4期416-424,共9页Journal of Zhejiang University:Agriculture and Life Sciences

基　　金：Projects supported by Zhejiang Provincial Natural Science Foundation of China(No.LR16H300001);National Natural Science Foundation of China(No.31670008)

摘　　要：棘白菌素类抗真菌药物米卡芬净和阿尼芬净的合成工艺中包括一个关键步骤:水解除去FR901379分子和棘白菌素B(echinocandin B,ECB)分子的脂肪酸侧链,形成环状的6元多肽核心。FR901379和ECB的水解可以分别被FR901379酰基酶和阿库来菌素A酰基酶(aculeacin A acylase,AAC)催化,因此,FR901379酰基酶和AAC的发掘、表征和生产在米卡芬净和阿尼芬净的工业生产上具有重要的应用价值。本研究首先筛选到了犹他游动放线菌SW1311,发现该菌株发酵液具有酰基酶活性,并摸索了不同发酵条件对酰基酶活性的影响。然后将犹他游动放线菌SW1311中的AAC基因克隆到改造过的质粒载体p IJ8660中,并将该质粒转化到天蓝色链霉菌(Streptomyces coelicolor)A3(2)中对AAC基因进行高表达,得到重组菌株sSCO-AAC。最后将sSCO-AAC生产的AAC活性和所需培养时间与犹他游动放线菌SW1311进行比较,表征了sSCO-AAC的发酵液水解FR901379的反应。结果表明,犹他游动放线菌SW1311中的酰基酶具有水解FR901379和青霉素V的酰基活性。用重组菌株sSCO-AAC生产的AAC活性比犹他游动放线菌SW1311的高4.6倍,且该重组菌株所需的培养时间比犹他游动放线菌SW1311缩短了30%。该结果不仅将ACC的应用范围从阿尼芬净合成拓宽到了米卡芬净合成,而且还揭示出天蓝色链霉菌A3(2)可以作为一个良好的AAC表达菌株。本研究对阿尼芬净和米卡芬净的工业化生产具有潜在的应用价值。Echinocandins are the first class of antifungals to target fungal cell wall.Two antifungals,including micafungin and anidulafungin,have been widely used in clinic.The synthesis of them includes an essential process:hydrolytic removal of fatty acyl side chains from FR901379and echinocandin B(ECB)to generate cyclic hexapeptide nuclei.In this study,Actinoplanes utahensis SW1311with acylase activity was isolated for both FR901379and penicillin V.Then a recombinant Streptomyces coelicolor A3(2)harboring aculeacin A acylase(AAC)gene from A.utahensis SW1311was constructed.The result showed that the AAC activity produced by the recombinant strain was4.6-fold higher than that of A.utahensis SW1311.Additionally,the fermentation time of the recombinant strain was30%shorter than that of A.utahensis SW1311.The results not only provide a new application of AAC for micafungin synthesis but also identify a new suitable host for AAC gene.

关 键 词：米卡芬净 FR901379 棘白菌素 抗真菌药物 阿库来菌素A酰基酶 犹他游动放线菌 

分 类 号：Q81[生物学—生物工程]