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Background  β-Mannanase catalyzes the cleavage of β-1,4-linked internal linkages of mannan backbone randomly to produce new chain ends. Alkaline and thermostable β-mannanases provide obvious advantages for many applications in biobleaching of pulp and paper, detergent industry, oil grilling operation and enzymatic production of mannooligosaccharides. However, only a few of them are commercially exploited as wild or recombinant enzymes, and none heterologous and secretory expression of alkaline β-mannanase in  Bacillus subtilis  expression system was reported. Alkaliphilic  Bacillus clausii  S10 showed high β-mannanase activity at alkaline condition. In this study, this β-mannanase was cloned, purified and characterized. The high-level secretory expression in  B. subtilis  was also studied.    Results  A thermo-alkaline β-mannanase (BcManA) gene encoding a 317-amino acid protein from alkaliphilic  Bacillus clausii  strain was cloned and expressed in  Escherichia coli . The purified mature BcManA exhibited maximum activity at pH 9.5 and 75 °C with good stability at pH 7.0–11.5 and below 80 °C. BcManA demonstrated high cleavage capability on polysaccharides containing β-1,4-mannosidic linkages, such as konjac glucomannan, locust bean gum, guar gum and sesbania gum. The highest specific activity of 2366.2 U mg −1  was observed on konjac glucomannan with the  K   m   and  k   cat   value of 0.62 g l −1  and 1238.9 s −1 , respectively. The hydrolysis products were mainly oligosaccharides with a higher degree of polymerization than biose. BcManA also cleaved manno-oligosaccharides with polymerization degree more than 3 without transglycosylation. Furthermore, six signal peptides and two strong promoters were used for efficiently secreted expression optimization in  B. subtilis  WB600 and the highest extracellular activity of 2374 U ml −1  with secretory rate of 98.5% was obtained using SP  lipA   and P43 after 72 h cultivation in 2 × SR medium. By medium optimization using cheap nitrogen and carbon source of peanut meal and glucose, the extracellular activity reached 6041 U ml −1  after 72 h cultivation with 6% inoculum size by shake flask fermentation.    Conclusions  The thermo-alkaline β-mannanase BcManA showed good thermal and pH stability and high catalytic efficiency towards konjac glucomannan and locust bean gum, which distinguished from other reported β-mannanases and was a promising thermo-alkaline β-mannanase for potential industrial application. The extracellular BcManA yield of 6041 U ml −1 , which was to date the highest reported yield by flask shake, was obtained in  B. subtilis  with constitutive expression vector. This is the first report for secretory expression of alkaline β-mannanase in  B. subtilis  protein expression system, which would significantly cut down the production cost of this enzyme. Also this research would be helpful for secretory expression of other β-mannanases in  B. subtilis .    Electronic supplementary material  The online version of this article (10.1186/s12934-018-0973-0) contains supplementary material, which is available to authorized users.

microbial cell factories

Characterization and high-efficiency secreted expression in Bacillus subtilis of a thermo-alkaline β-mannanase from an alkaliphilic Bacillus clausii strain S10