Secretion, purification, and characterization of a recombinant Aspergillus oryzae tannase in Pichia pastoris.
Tannase (tannin acyl hydrolase) is an industrially important enzyme produced by a large number of fungi, which hydrolyzes the ester and depside bonds of gallotannins and gallic acid esters. In the present work, a tannase from Aspergillus oryzae has been cloned and expressed in Pichia pastoris. The catalytic activity of the recombinant enzyme was assayed. A secretory form of enzyme was made with the aid of Saccharomyces cerevisiae alpha-factor, and a simple procedure purification protocol yielded tannase in pure form. The productivity of secreted tannase achieved 7000 IU/L by fed-batch culture. Recombinant tannase had a molecular mass of 90 kDa, which consisted of two kinds of subunits linked by a disulfide bond(s). Our study is the first report on the heterologous expression of tannase suggesting that the P. pastoris system represents an attractive means of generating large quantities of tannase for both research and industrial purpose.
Zhong
Xiaofen
X
The Open Laboratory for Marine Functional Genomics of State High-Tech Development, Key Laboratory of Genetic Engineering of MOE, Department of Biochemistry, College of Life Sciences, Sun Yat-Sen University, Guangzhou 510275, People's Republic of China.
Peng
Lisheng
L
Zheng
Suilan
S
Sun
Zhizhi
Z
Ren
Yufeng
Y
Dong
Meiling
M
Xu
Anlong
A
D001236:Aspergillus oryzae / Q000201:enzymology* / Q000235:genetics; D002265:Carboxylic Ester Hydrolases / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D004220:Disulfides / Q000737:chemistry; D005656:Fungal Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic; D015966:Gene Expression Regulation, Fungal; D000072235:Mating Factor; D008970:Molecular Weight; D010455:Peptides / Q000737:chemistry / Q000235:genetics; D010843:Pichia / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000737:chemistry / Q000235:genetics
D004220:Disulfides; D005656:Fungal Proteins; D010455:Peptides; D000072235:Mating Factor; D002265:Carboxylic Ester Hydrolases; C022711:tannase