title: High-level expression and characterization of a thermophilic β-mannanase from Aspergillus niger in Pichia pastoris.
abstract: OBJECTIVE
  A novel, high-level expression, thermostable mannan endo-1,4-beta-mannosidase is urgently needed for industrial applications.
  
  
  RESULTS
  The mannan endo-1,4-β-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 ~3.5 mg/ml and the enzyme activity was 1612 U/ml. The enzyme exhibited a maximum activity of 3049 U/ml at 80 °C and retained 60% enzyme activity at 80 °C for 2 h. The pH optimum was 4.5 and the enzyme was stable over the pH range 1.5-11.
  
  
  CONCLUSIONS
  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.
authors.affiliation: Hubei Collaborative Innovation Center for Green Transformation of Bio-resources, Hubei Key Laboratory of Industrial Biotechnology, College of Life Sciences, Hubei University, Wuhan, 430062, People's Republic of China.
authors.forename: Shi
authors.initials: S
authors.lastname: Yu
authors.forename: Zhezhe
authors.initials: Z
authors.lastname: Li
authors.forename: Yaping
authors.initials: Y
authors.lastname: Wang
authors.forename: Wanping
authors.initials: W
authors.lastname: Chen
authors.forename: Lin
authors.initials: L
authors.lastname: Fu
authors.forename: Wei
authors.initials: W
authors.lastname: Tang
authors.forename: Chen
authors.initials: C
authors.lastname: Chen
authors.forename: Yunyun
authors.initials: Y
authors.lastname: Liu
authors.forename: Xue
authors.initials: X
authors.lastname: Zhang
authors.forename: Lixin
authors.initials: L
authors.lastname: Ma
mesh_terms: D001234:Aspergillus niger / Q000201:enzymology* / Q000235:genetics; D003001:Cloning, Molecular; D004795:Enzyme Stability; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism*; D006031:Glycosylation; D006863:Hydrogen-Ion Concentration; D010843:Pichia / Q000235:genetics* / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D013696:Temperature; D044902:beta-Mannosidase / Q000235:genetics / Q000378:metabolism*
chemical_list: D005656:Fungal Proteins; D011994:Recombinant Proteins; D044902:beta-Mannosidase