N-乙酰基色胺的异源生物合成和抑藻活性研究  

Heterogenous biosynthesis and algal inhibition activity of N-acetyltryptamine

作　　者：张胜男 朱春苗 江婧娴 马丽艳[2] 张风丽 Zhang Sheng-nan;Zhu Chun-miao;Jiang Jing-xian;Ma Li-yan;Zhang Feng-li(State Key Laboratory of Microbial Metabolism,School of Life Sciences and Biotechnology,Shanghai Jiao Tong University,Shanghai 200240;East China Sea Fisheries Research Institute,Shanghai 200090)

机构地区：[1]上海交通大学生命科学技术学院微生物代谢国家重点实验室,上海200240 [2]中国水产科学研究院东海水产研究所,上海200090

出　　处：《中国抗生素杂志》2020年第2期137-144,共8页Chinese Journal of Antibiotics

基　　金：国家重点研发计划(No.2018YFC0310900)。

摘　　要：目的 用Streptomyces lividans GX28异源合成N-乙酰基色胺,并研究N-乙酰基色胺的抗赤潮藻活性。方法 前期研究发现Bacillus atrophaeus C89芳香族氨基酸脱羧酶(aromatic l-amino acid decarboxylase, AADC)能将L-色氨酸脱羧成L-色胺。氨基酸多序列比对发现S. lividans GX28基因组中存在使色胺乙酰化的酶。通过克隆、重组将aadc基因转入S. lividans GX28中构建重组菌株,HPLC检测目标产物,NMR鉴定化合物的结构。检测化合物对3种赤潮微藻的毒性。结果 S. lividans GX28中的蛋白WP_015609847.1含有芳烷基胺N-乙酰转移酶(alkyl amine N-acetyltransferase, AANAT)的底物结合位点(P_(64)和F_(188))和催化位点(Y_(168)),推测其为AANAT。宿主的发酵产物中分离得到N-乙酰基色胺。N-乙酰基色胺对墨西哥原甲藻的生长具有较强的抑制作用(IC_(50)=9.56mg/L)。结论 推测aadc基因在链霉菌中异源表达。重组链霉菌中检测到N-乙酰基色胺,并对N-乙酰基色胺的生物合成途径进行推测。N-乙酰基色胺对P. mexicanum的生长具有显著的抑制作用。本研究为抑藻化合物的生物合成研究奠定了基础。Objective Streptomyces lividans GX28 was used to heterologous synthesis of N-acetyltryptamine,and the activity of N-acetyltryptamine against red tide algae was tested.Methods Previous studies showed that aromatic L-amino acid decarboxylase(AADC)from Bacillus atrophaeus C89,can decarboxylatel-tryptophan to L-tryptamine.N-acetyltransferase amino acid sequences from S.lividans GX28 and other organisms were analyzed using the amino acid multiple sequence alignment.Then,aadc gene from B.atrophaeus C89 was cloned and transformed into S.lividans GX28 HPLC was used to detect the target product from recombinant strain.Finally,the structure of the target product was determined by NMR and the toxicity of the compound against Prorocentrum lima,Alexandrium tamarense,and Prorocentrum mexicanum was tested.Results The protein WP_015609847.1 of S.lividans GX28 contains substrate binding sites(P64 and F188)and catalytic site(Y168)of alkyl amine N-acetyltransferase(AANAT),which was presumed to be members of the AANAT family.N-acetyltryptamine was isolated from the fermentation products of S.lividans GX28.N-acetyltryptamine showed a significant inhibitory effect on P.mexicanum(IC50=9.56mg/L).Conclusion The aadc gene was heterologously expressed in S.lividans GX28.The target product N-acetyltryptamine was detected from recombinant Streptomyces strain.The biosynthesis pathway of N-acetyltryptamine was proposed.N-acetyltryptamine exhibited high algal inhibitory activity against P.mexicanum.The study laid a foundation for the biosynthesis of algae-inhibiting compounds.

关 键 词：N-乙酰基色胺 AANAT aadc基因 异源表达 抑藻剂 

分 类 号：R9[医药卫生—药学]