多酶级联反应合成能够促进肠道益生菌生长的纤维寡糖  

Synthesis of cello-oligosaccharides which promotes the growth of intestinal probiotics by multi-enzyme cascade reaction

作　　者：郑棚 王雷 胡美荣[2] 魏华[1] 陶勇[2,3] ZHENG Peng;WANG Lei;HU Meirong;WEI Hua;TAO Yong(Jiangxi-OAI Joint Research Institute,Nanchang University,Nanchang 330047,Jiangxi,China;CAS Key Laboratory of Microbial Physiological and Metabolic Engineering,Institute of Microbiology,Chinese Academy of Sciences,Beijing 100101,China;University of Chinese Academy of Sciences,Beijing 100049,China)

机构地区：[1]南昌大学中德联合研究院,江西南昌330047 [2]中国科学院微生物研究所中国科学院微生物生理与代谢工程重点实验室,北京100101 [3]中国科学院大学,北京100049

出　　处：《生物工程学报》2023年第8期3406-3420,共15页Chinese Journal of Biotechnology

基　　金：国家重点研发计划(2017YFD0502102-3)。

摘　　要：聚合度2–6的可溶性纤维寡糖是一种具有多种生物功能的低聚糖,它能够促进双歧杆菌(Bifidobacteria)、副干酪乳杆菌(Lactobacillus paracei)等肠道益生菌的增殖,因此对人体肠道微生态具有调节作用。本研究通过在大肠杆菌中表达纤维寡糖磷酸化酶(cellodextrin phosphorylase,CDP),构建Cc 01菌株,并与之前构建的COS 01菌株联合使用,建立了基于COS 01、Cc 01的三酶级联反应催化底物葡萄糖和蔗糖合成纤维寡糖反应体系。经过优化后,最终可溶性纤维寡糖的产量达到97g/L,纯度约为97%,其中含有纤维二糖(16.8wt%)、纤维三糖(49.8wt%)、纤维四糖(16.4 wt%)、纤维五糖(11.5 wt%)和纤维六糖(5.5 wt%)。在纤维寡糖对益生菌株生长促进作用的测试中,以菊粉、低聚木糖、低聚果糖为基准,干酪乳杆菌(WSH004)、副干酪乳杆菌(WSH005)以及嗜酸乳杆菌(WSH 006)利用纤维寡糖(聚合度2–6)为碳源进行生长后,益生菌的生物量(OD600)相比对照增加约2倍。该研究证明了三酶级联反应能够高效合成纤维寡糖,并表明聚合度2–6的纤维寡糖是一类具有促进肠道微生物增殖的功能性碳水化合物。Soluble cello-oligosaccharide with 2–6 oligosaccharide units is a kind of oligosaccharide with various biological functions,which can promote the proliferation of intestinal probiotics such as Bifidobacteria and Lactobacillus paracei.Therefore,it has a regulatory effect on human intestinal microbiota.In this study,a Cc 01 strain was constructed by expressing cellodextrin phosphorylase(CDP)in Escherichia coli.By combining with a previously constructed COS 01 strain,a three-enzyme cascade reaction system based on strains COS 01 and Cc 01 was developed,which can convert glucose and sucrose into cello-oligosaccharide.After optimization,the final titer of soluble cello-oligosaccharides with 2–6 oligosaccharide units reached 97 g/L,with a purity of about 97%.It contained cellobiose(16.8 wt%),cellotriose(49.8 wt%),cellotetrose(16.4 wt%),cellopentaose(11.5 wt%)and cellohexose(5.5 wt%).When using inulin,xylo-oligosaccharide and fructooligosaccharide as the control substrate,the biomass(OD600)of Lactobacillus casei(WSH 004),Lactobacillus paracei(WSH 005)and Lactobacillus acidophilus(WSH 006)on cello-oligosaccharides was about 2 folds higher than that of the control.This study demonstrated the efficient synthesis of cello-oligosaccharides by a three-enzyme cascade reaction and demonstrated that the synthesized cello-oligosaccharides was capable of promoting intestinal microbial proliferation.

关 键 词：纤维寡糖 全细胞催化 磷酸化酶 功能性碳水化合物 肠道益生菌 

分 类 号：Q789[生物学—分子生物学]