Chinese Academy of Sciences Key Laboratory of Microbial Physiological and Metabolic Engineering, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China.
College of Life Science, University of Chinese Academy of Sciences, Beijing 100049, China.
J Ind Microbiol Biotechnol. 2022 May 25;49(3). doi: 10.1093/jimb/kuac008.
Cellobiose, a natural disaccharide, attracts extensive attention as a potential functional food/feed additive. In this study, we present an inorganic phosphate (Pi) self-sufficient biotransformation system to produce cellobiose by co-expressing sucrose phosphorylase (SP) and cellobiose phosphorylase (CBP). The Bifidobacterium adolescentis SP (BASP) and Cellvibrio gilvus CBP (CGCBP) were co-expressed in Escherichia coli. Escherichia coli cells containing BASP and CGCBP were used as whole-cell catalysts to convert sucrose and glucose to cellobiose. The effects of reaction pH, temperature, Pi concentration, and substrate concentration were investigated. In the optimum biotransformation conditions, 800 mM cellobiose was produced from 1.0 M sucrose, 1.0 M glucose, and 50 mM Pi, within 12 hr. The by-product fructose and residual substrate (sucrose and glucose) were efficiently removed by treatment with yeast, to help purify the product cellobiose. The wider applicability of this Pi self-sufficiency strategy was demonstrated in the production of laminaribiose by co-expressing SP and laminaribiose phosphorylase. This study suggests that the Pi self-sufficiency strategy through co-expressing two phosphorylases has the advantage of great flexibility for enhanced production of cellobiose (or laminaribiose).
纤维二糖是一种天然二糖，作为一种有潜力的功能性食品/饲料添加剂而受到广泛关注。在本研究中，我们提出了一种无机磷酸盐（Pi）自足的生物转化系统，通过共表达蔗糖磷酸化酶（SP）和纤维二糖磷酸化酶（CBP）来生产纤维二糖。双歧杆菌 SP（BASP）和吉尔沃氏梭菌 CBP（CGCBP）在大肠杆菌中共同表达。含有 BASP 和 CGCBP 的大肠杆菌细胞被用作全细胞催化剂，将蔗糖和葡萄糖转化为纤维二糖。考察了反应 pH 值、温度、Pi 浓度和底物浓度的影响。在最佳生物转化条件下，从 1.0 M 蔗糖、1.0 M 葡萄糖和 50 mM Pi 中，在 12 小时内生产了 800 mM 的纤维二糖。酵母处理有效地去除了副产物果糖和残留的底物（蔗糖和葡萄糖），有助于纯化产物纤维二糖。通过共表达 SP 和纤维二糖磷酸化酶生产纤维三糖，证明了这种 Pi 自足策略的更广泛适用性。本研究表明，通过共表达两种磷酸化酶的 Pi 自足策略在增强纤维二糖（或纤维三糖）生产方面具有很大的灵活性优势。
