The mutagenic effects of the ultraviolet ray, diethyl sulfate's single factor and ultraviolet-diethyl sulfate compound mutagenesis on Bacillus subtilis HD-1 (54.6 U/mL) were studied. Results of enzyme activity showed that ultraviolet-diethyl sulfate compound mutagenesis was better. A mutant strain, named Bacillus subtilis UD-20 with high-level β-mannanase production was obtained after ultraviolet-diethyl sulfate compound mutagenesis from Bacillus subtilis HD-1 which was an endophytic bacteria screened from soybean seeds. The β-mannanase activity of Bacillus subtilis UD-20 increased 63.9% compared with that of the original strain Bacillus subtilis HD-1 (54.6 U/ml) and was stable during the generations. The optimal temperature for β-mannanase activity was 50 ℃ and the enzyme was stable up to 50 ℃. The optimal pH for enzyme activity was 7.0 and the enzyme demonstrated broad pH stability within a pH range of 5.0~8.0. The β-mannanase activity was activated by Fe2+ (113.2%) and Co2+ (131.7%), but strongly inhibited by Mn2+ (55.6%). The β-mannanase stability was increased by K+, Ca2+, Ba2+, among which Co2+ (150%) showed the best effect on the enzyme stability.
[1] Dhawan S, Kaur J. Microbial mannanases: an overview of production and applications [J]. Critical Reviews in Biotechnology, 2007, 27: 197-216
[2] Cai H, Shi P, Huang H, et al. An acidic β-mannanase from Penicillium sp. C6: gene cloning and over-expression in Pichia pastoris [J]. World Journal of Microbiology and Biotechnology, 2011, 27(12): 2813-2819
[5] Promputtha I, Hyde K D, McKenzie E H C, et al. Can leaf degrading enzymes provide evidence that endophytic fungi becoming saprobes [J]. Fungal Diversity, 2010, 41(1): 89-99
[6] Cho K M, Hong S Y, Lee S M, et al. A cel44C-man26A gene of endophytic PaeniBacillus polymyxa GS01 has multi- glycosyl hydrolases in two catalytic domains [J]. Appled Microbiology and Biotechnology, 2006, 73(3): 618-630
[7] Kima D Y, Hama S J, Leea H J, et al. A highly active endo-β-1, 4-mannanase produced by Cellulosimicrobium sp. strain HY-13, a hemicellulolytic bacterium in the gut of Eisenia fetida [J]. Enzyme and Microbial Technology, 2011, 48(4-5,7): 365-370
[12] Summpunn P, Chaijan S, Isarangkul D, et al. Characterization, gene cloning, and heterogeneous expression of β-mannanase from thermophilic Bacillus subtilis [J]. The Journal of Microbiology, 2011, 49(1): 86-93
