Journal
BIOTECHNOLOGY LETTERS
Volume 37, Issue 9, Pages 1853-1859Publisher
SPRINGER
DOI: 10.1007/s10529-015-1848-7
Keywords
High-density fermentation; High-level expression; Industry; Mannanase; Oligosaccharides; Pichia pastoris; Thermophilic
Categories
Funding
- National Natural Science Foundation of China [31100057, 31172320, 31340036]
- National 973 Program of China [2013CB910801]
- Nature Science Fund for Creative Research Groups of Hubei Province of China [2012FFA034]
- Foundation for High and New Technology Industrial Innovative Research Groups of the Wuhan Science and Technology Bureau's Department [2014070504020239]
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A novel, high-level expression, thermostable mannan endo-1,4-beta-mannosidase is urgently needed for industrial applications. The mannan endo-1,4-beta-mannosidase gene (MAN) from Aspergillus niger CBS 513.88 was optimized based on the codon usage bias in Pichia pastoris and synthesized by overlapping PCR to produce MAN-P. It was expressed in P. pastoris GS115 from a constitutive expression vector pHBM-905 M. MAN-P reached 594 mg/l in shake-flasks after 192 h induction. On production in a 5 l fermenter, the yield of MAN-P reached similar to 3.5 mg/ml and the enzyme activity was 1612 U/ml. The enzyme exhibited a maximum activity of 3049 U/ml at 80 A degrees C and retained 60 % enzyme activity at 80 A degrees C for 2 h. The pH optimum was 4.5 and the enzyme was stable over the pH range 1.5-11. The thermostability of MAN-P is higher than other known fungal mannanases and the expression and thermophilic properties make MAN-P useful for industrial applications.
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