SDR基因在嗜热杜邦菌Thermolides生物合成中的作用  

Functions of SDR in thermolide biosynthesis by Thermomyces dupontii

作　　者：杜远江 黄为平 张克勤[1] 牛雪梅[1] DU Yuanjiang;HUANG Weiping;ZHANG Keqin;NIU Xuemei(State Key Laboratory for Conservation and Utilization of Bio-Resources in Yunnan,Yunnan University,Kunming 650091,China)

机构地区：[1]云南大学省部共建云南生物资源保护与利用国家重点实验室,昆明650091

出　　处：《应用与环境生物学报》2021年第6期1415-1426,共12页Chinese Journal of Applied and Environmental Biology

基　　金：国家自然科学基金项目(21977086,21867018)资助。

摘　　要：对嗜热杜邦菌Thermomyces dupontii中负责生物合成PKS-NRPS大环内酯Thermolides的关键基因进行异源宿主表达的前期研究发现,短链脱氢酶(short chain dehydrogenase,SDR)基因负责Thermolides中6-羰基还原成羟基,并是生物合成Thermolides A-C的最后反应步骤.对原宿主嗜热杜邦菌中基因功能缺失进行研究,发现乙酰基转移酶(acetyltransferase,AT)基因负责的Thermolides乙酰化修饰是合成Thermolides A-C最后反应步骤.因此进一步对嗜热杜邦菌突变菌株ΔSDR和ΔAT进行发酵,并比较分析这两株突变菌株代谢图谱中的差异化合物.在ΔSDR中,未检测到异源宿主表达研究中Thermolides的6位羰基的中间体及其衍生物,也未检测到其他可能的所有生物合成Thermolides的羰基中间体及其衍生物.研究结果表明SDR基因不仅负责Thermolides中6位酮基还原为羟基,也可能参与了Thermolides骨架的形成;这为解析类似的SDR基因的原有功能提供了一定思路,同时表明通过异源宿主表达和本宿主基因编辑来研究生物合成基因的功能会产生差异.Previous studies have reported that four key genes are responsible for the biosynthesis of potent nematicidal PKS-NRPS thermolides in Thermomyces dupontii. Through the heterologous expression method,a short chain dehydrogenase gene(SDR) was found to be involved in the reduction of 6-ketone in thermolides,which is also the last step in thermolide A-C biosynthesis. In this study, mutation analysis of T. dupontii identified an acetyltransferase gene(AT) responsible for the acetylation of thermolide D to A-C that was found to be involved in the last step in thermolide A-C biosynthesis. In this study, two T. dupontii mutants, ΔSDR and ΔAT,were fermented, and the extracts of the culture broths were carefully analyzed using HPLC-MS/MS. However, no 6-ketone thermolide intermediates and related analogs were detected in the metabolic profile of mutant ΔSDR,and other carbonyl thermolide intermediates and related analogs were also not detected. Our results indicate that SDR might also be involved in the ring formation of thermolides rather than only in the reduction of 6-ketone. Our results also suggest differences in the functions of biosynthetic genes in heterologous and native hosts.

关 键 词：杜邦嗜热菌 Thermolides 短链脱氢酶基因(SDR) 异源表达 基因功能 

分 类 号：Q78[生物学—分子生物学]