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Fusion and secretory expression of an exo‐inulinase and a d ‐allulose 3‐epimerase to produce d ‐allulose syrup from inulin
Author(s) -
Huang Yueyuan,
Li Liangfei,
Chi Yaowei,
Sha Yuanyuan,
Wang Rui,
Xu Zheng,
Xu Xiaoqi,
Li Sha,
Gao Zhen,
Xu Hong
Publication year - 2020
Publication title -
journal of the science of food and agriculture
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.782
H-Index - 142
eISSN - 1097-0010
pISSN - 0022-5142
DOI - 10.1002/jsfa.10682
Subject(s) - inulinase , inulin , chemistry , fructose , jerusalem artichoke , hydrolysis , enzyme , biochemistry , food science , yield (engineering) , materials science , metallurgy
BACKGROUND This study developed a feasible catalytic method for d ‐allulose syrup production using a fusion enzyme, either in free or immobilized form, through hydrolysis of inulin extracted from Jerusalem artichoke tubers. RESULTS d ‐Allulose 3‐epimerase (DAE) was actively expressed in secretory form by fusing with the extracellular exo‐inulinase CSCA in Escherichia coli BL21 (DE3). The best linker ligating the two enzymes was a flexible peptide containing 12 residues (GSAGSAAGSGEF). At 55 °C and pH 8.0, and as with the addition of 1 mmol L −1 Mn 2+ , the CSCA‐linkerE‐DAE fusion enzyme obtained through high cell‐density cultivation displayed a maximal exo‐inulinase activity of 21.8 U mg −1 and resulted in a yield of 6.3 g L −1 d ‐allulose and 39.2 g L −1 d ‐fructose using 60 g L −1 inulin as the raw material. Catechol‐modified alginate with titanium ions (Alg(Ti)PDA) was found to be a promising immobilization material for the fusion enzyme. After conversion for 8 days, the Alg(Ti)PDA‐immobilized CSCA‐linkerE‐DAE (8 U g −1 ) completed 24 reaction cycles and retained over 80% of its original activity. Each reaction obtained an average of 19.8 g L −1 d ‐allulose and 32.7 g L −1 D‐fructose from 60 g L −1 inulin. CONCLUSION This study shed light on a feasible and cost‐effective approach for the production of syrup containing d ‐allulose and D‐fructose with inulin as the raw material via the use of a CSCA and DAE fusion enzyme. This syrup is of added value as a functional sweetener. © 2020 Society of Chemical Industry