Journal
PROTEIN EXPRESSION AND PURIFICATION
Volume 139, Issue -, Pages 29-35Publisher
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/j.pep.2017.07.005
Keywords
beta-mannanase; Bacillus circulans NT 6.7; pSIP403; Lactobacillus plantarum WCFS1; Manno-oligosaccharides
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Funding
- Center for Advanced Studies for Agriculture and Food, Institute for Advanced Studies, Kasetsart University under the Higher Education Research Promotion
- National Research University Project of Thailand, Office of the Higher Education Commission, Ministry of Education, Thailand [CASAF161]
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The beta-mannanase gene of Bacillus circulans NT 6.7 was successfully cloned in Lactobacillus plantarum WCFS1 using the pSIP403 expression vector and secreted to the supernatant rather than accumulated in the cells. The highest activity was achieved by controlling the pH at 6 during cultivation. Maximum mannanase activities detected in the supernatant and cell-free extract of 200 ml MRS broth were 8.2 and 0.86 U/ml, respectively. Enzyme activity in the supernatant increased to 27 U/ml by fermentation in a 5-L bioreactor with automatic pH control. The optimum temperature of recombinant beta-mannanase was 50 degrees C and stable between 30 and 50 degrees C. The optimum pH was 6 with stability in the range 5-7. Enzyme activity slightly increased with Co2+ but was strongly inhibited by EDTA. The enzyme exhibited high specificity to galactomannan substrates. The main products of copra meal and locust bean gum hydrolysis were manno-oligosaccharides. Therefore, recombinant beta-mannanase produced from a food grade host, L. plantarum WCFS1, showed potential for use in manno-oligosaccharides production and other food-related applications. (C) 2017 Elsevier Inc. All rights reserved.
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