TY - JOUR
T1 - Efficient production of recombinant tannase in Aspergillus oryzae using an improved glucoamylase gene promoter
AU - Ichikawa, Kyotaro
AU - Shiono, Yoshihito
AU - Shintani, Tomoko
AU - Watanabe, Akira
AU - Kanzaki, Hiroshi
AU - Gomi, Katsuya
AU - Koseki, Takuya
N1 - Funding Information:
This study was financially supported in part by the Strategic Core Technology Advancement Program of the Ministry of Economy, Trade and Industry, Japan. The authors declare that they have no conflicts of interest to this work.
Publisher Copyright:
© 2019 The Society for Biotechnology, Japan
PY - 2020/2
Y1 - 2020/2
N2 - A tannase-encoding gene, AotanB, from Aspergillus oryzae RIB40 was overexpressed in A. oryzae AOK11 niaD-deficient mutant derived from an industrial strain under the control of an improved glucoamylase gene promoter PglaA142. The recombinant tannase, designated as rAoTanBO, was produced efficiently as an active extracellular enzyme. Purified rAoTanBO showed a smeared band with a molecular mass of approximately 80–100 kDa on sodium dodecyl sulfate polyacrylamide gel electrophoresis. The rAoTanBO had a molecular mass of 65 kDa, after treatment with endo-β-N-acetylglucosaminidase H. Purified rAoTanBO exhibited maximum activity at 30–35°C and pH 6.0. The tannase activity of purified rAoTanBO towards natural and artificial substrates was 2–8 folds higher than that of the recombinant enzyme produced by Pichia pastoris, designated as rAoTanBP. N-terminus of the mature rAoTanBP had six more amino acids than the N-terminus of the mature rAoTanBO. Kinetic analyses showed that rAoTanBO had higher catalytic efficiency (kcat/Km) than rAoTanBP. rAoTanBO was stable up to 60°C and higher thermostability than rAoTanBP. N-linked oligosaccharides had no effect on the activity and stability of rAoTanBO and rAoTanBP.
AB - A tannase-encoding gene, AotanB, from Aspergillus oryzae RIB40 was overexpressed in A. oryzae AOK11 niaD-deficient mutant derived from an industrial strain under the control of an improved glucoamylase gene promoter PglaA142. The recombinant tannase, designated as rAoTanBO, was produced efficiently as an active extracellular enzyme. Purified rAoTanBO showed a smeared band with a molecular mass of approximately 80–100 kDa on sodium dodecyl sulfate polyacrylamide gel electrophoresis. The rAoTanBO had a molecular mass of 65 kDa, after treatment with endo-β-N-acetylglucosaminidase H. Purified rAoTanBO exhibited maximum activity at 30–35°C and pH 6.0. The tannase activity of purified rAoTanBO towards natural and artificial substrates was 2–8 folds higher than that of the recombinant enzyme produced by Pichia pastoris, designated as rAoTanBP. N-terminus of the mature rAoTanBP had six more amino acids than the N-terminus of the mature rAoTanBO. Kinetic analyses showed that rAoTanBO had higher catalytic efficiency (kcat/Km) than rAoTanBP. rAoTanBO was stable up to 60°C and higher thermostability than rAoTanBP. N-linked oligosaccharides had no effect on the activity and stability of rAoTanBO and rAoTanBP.
KW - AotanB gene
KW - Aspergillus oryzae
KW - Efficient production
KW - Over-expression
KW - Tannase
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U2 - 10.1016/j.jbiosc.2019.08.002
DO - 10.1016/j.jbiosc.2019.08.002
M3 - Article
C2 - 31492608
AN - SCOPUS:85071594597
SN - 1389-1723
VL - 129
SP - 150
EP - 154
JO - Journal of Bioscience and Bioengineering
JF - Journal of Bioscience and Bioengineering
IS - 2
ER -