Alkaline mannanase from a novel species of alkaliphilic Bacillus

• Published in: Journal of Applied Glycoscience Vol. 51; no. 3; pp. 229 - 236
• Main Authors: Takeda, N. (Toyo Univ., Tokyo (Japan)), Hirasawa, K, Uchimura, K, Nogi, Y, Hatada, Y, Akita, M, Usami, R, Yoshida, Y, Grant, W.D, Ito, S, Horikoshi, K
• Format: Journal Article
• Language: English; Japanese
• Published: The Japanese Society of Applied Glycoscience 2004
• Subjects: alkaline enzyme; alkaliphile; BACILLUS; GENES; glycoside hydrolase family 5; HYDROLASES; mannanase; MANNANS; NUCLEOTIDE SEQUENCE
• ISSN: 1344-7882; 1880-7291
• DOI: 10.5458/jag.51.229

A novel species of alkaliphilic Bacillus (JAMB-602), isolated from a soil sample, was found to exoproduce an alkaline mannanase. The mannanase gene was cloned by the shotgun method and sequenced. Its open reading frame encodes a protein of 490 amino acids with a calculated molecular mass of 53,763 Da. The deduced amino acid sequence showed similarity to those of known mannanases in glycoside hydrolase family 5, with only 32-58% identity. The recombinant mature enzyme with a molecular mass of 50 kDa was hyperexoproduced with Bacillus subtilis as the host, corresponding to a level of approximately 2 g/L. The homogeneously purified enzyme had a molecular mass of 50 kDa, an optimal temperature of 65 deg C, and an optimal pH of around 9. It was thermostable with a half-life of 2 h at 55 deg C. N -Bromosuccinimide abolished the enzymatic activity. The pattern of mannan hydrolysis showed that the enzyme is an endo-type mannanase.