重组恶臭假单胞菌转化乙二醇-乙酸共底物合成单鼠李糖脂  

Production of mono-rhamnolipid from co-substrate of ethylene glycol and acetic acid by recombinant Pseudomonas putida

作　　者：辛开元 何美霖 罗云鹏 刘豪杰 周杰 钱秀娟[1] 董维亮 XIN Kaiyuan;HE Meilin;LUO Yunpeng;LIU Haojie;ZHOU Jie;QIAN Xiujuan;DONG Weiliang(Key Laboratory for Waste Plastics Biocatalytic Degradation and Recycling,College of Biotechnology and Pharmaceutical Engineering,Nanjing Tech University,Nanjing 211800,China;2011 College,Nanjing Tech University,Nanjing 211800,China;State Key Laboratory of Materials-Oriented Chemical Engineering,Nanjing Tech University,Nanjing 211800,China)

机构地区：[1]南京工业大学,生物与制药工程学院,废塑料生物催化解聚与循环利用重点实验室,江苏南京211800 [2]南京工业大学2011学院,江苏南京211800 [3]南京工业大学材料化学工程国家重点实验室,江苏南京211800

出　　处：《生物加工过程》2022年第4期457-464,共8页Chinese Journal of Bioprocess Engineering

基　　金：国家自然科学基金(31961133017、21978129);江苏省优秀青年基金(BK20211591);江苏省科协青年科技人才托举工程(TJ-2021-092);江苏省先进生物制造创新中心自主课题(XTD2205、XTD2203)。

摘　　要：乙二醇(ethylene glycol,EG)是聚对苯二甲酸乙二醇酯(PET)等塑料解聚后的单体产物之一,开发EG为原料的生物转化系统对于解决发酵行业原料替代和塑料资源化利用均具有重要意义。将EG与廉价碳源——乙酸(AC)进行共底物发酵,通过底物浓度优化获得最佳EG与乙酸配比为3∶4,可实现底物的完全消耗,解决了因乙酰辅酶A供给不足导致Pseudomonas putida KT2440难以利用EG的问题;进一步分别组装Pseudomonas aeruginosa PAO1和Pseudomonas aeruginosa KT1115来源的单鼠李糖脂(mono-RL)合成基因线路rhlIRBA,获得mono-RL合成恶臭假单胞菌重组菌株P.putida KT2441和P.putida KT2442。P.putida KT2442表现出更强的mono-RL合成能力,在50 mL摇瓶发酵体系中,P.putida KT2442利用EG和乙酸共底物合成mono-RL产量为0.46 g/L,得率为0.055 g/g;薄层色谱和质谱结果表明合成的mono-RL主要为Rha-C10-C10结构,分子量为503。Ethylene glycol(EG)is one of the main monomers generated from polyethylene terephthalate(PET),therefore,the development of EG biotransformation system shows great significance to promote both low-value fermenting substrate substitution and PET resource recycling.In this study,a co-substrate fermentation system using EG and acetic acid as carbon source was constructed.The co-substrate system promoted EG metabolism in Pseudomonas putida KT2440 by providing sufficient acetyl-CoA,and the optimal ratio of EG and acetic acid at 3∶4 could guarantee the complete consumption of all carbon source.Further,the key gene cluster of rhlIRBA responsible for mono-rhamnolipid(mono-RL)synthesis from Pseudomonas aeruginosa PAO1 and Pseudomonas aeruginosa KT1115 was cloned and overexpressed in P.putida KT2440,respectively.The recombinant P.putida KT2442 performed better mono-RL synthesis capability,as 0.46 g/L of mono-RL with 0.055 g/g of yield was obtained in flask fermentation system using EG and acetic acid as co-carbon source.Thin layer chromatography(TLC)and mass spectrum(MS)showed the synthesized mono-RL was mainly Rha-C10-C10 with a molecular weight of 503.

关 键 词：乙二醇 乙酸 塑料解聚物 共底物 单鼠李糖脂 恶臭假单胞菌 

分 类 号：Q815[生物学—生物工程]