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Journal of Zhejiang University SCIENCE B 2008 Vol.9 No.7 P.536~545

http://doi.org/10.1631/jzus.B0720006


Expression, purification and characterization of a phyAm-phyCs fusion phytase


Author(s):  Li-kou ZOU, Hong-ning WANG, Xin PAN, Guo-bao TIAN, Zi-wen XIE, Qi WU, Hui CHEN, Tao XIE, Zhi-rong YANG

Affiliation(s):  College of Life Science, Bioengineering Research Center for Animal Disease Prevention and Control, Sichuan University, Chengdu 610065, China; more

Corresponding email(s):   zoulkcn@hotmail.com, whongning@163.com

Key Words:  Expression, PhyAm, PhyCs, Fusion phytase, Pichia pastoris


Li-kou ZOU, Hong-ning WANG, Xin PAN, Guo-bao TIAN, Zi-wen XIE, Qi WU, Hui CHEN, Tao XIE, Zhi-rong YANG. Expression, purification and characterization of a phyAm-phyCs fusion phytase[J]. Journal of Zhejiang University Science B, 2008, 9(7): 536~545.

@article{title="Expression, purification and characterization of a phyAm-phyCs fusion phytase",
author="Li-kou ZOU, Hong-ning WANG, Xin PAN, Guo-bao TIAN, Zi-wen XIE, Qi WU, Hui CHEN, Tao XIE, Zhi-rong YANG",
journal="Journal of Zhejiang University Science B",
volume="9",
number="7",
pages="536~545",
year="2008",
publisher="Zhejiang University Press & Springer",
doi="10.1631/jzus.B0720006"
}

%0 Journal Article
%T Expression, purification and characterization of a phyAm-phyCs fusion phytase
%A Li-kou ZOU
%A Hong-ning WANG
%A Xin PAN
%A Guo-bao TIAN
%A Zi-wen XIE
%A Qi WU
%A Hui CHEN
%A Tao XIE
%A Zhi-rong YANG
%J Journal of Zhejiang University SCIENCE B
%V 9
%N 7
%P 536~545
%@ 1673-1581
%D 2008
%I Zhejiang University Press & Springer
%DOI 10.1631/jzus.B0720006

TY - JOUR
T1 - Expression, purification and characterization of a phyAm-phyCs fusion phytase
A1 - Li-kou ZOU
A1 - Hong-ning WANG
A1 - Xin PAN
A1 - Guo-bao TIAN
A1 - Zi-wen XIE
A1 - Qi WU
A1 - Hui CHEN
A1 - Tao XIE
A1 - Zhi-rong YANG
J0 - Journal of Zhejiang University Science B
VL - 9
IS - 7
SP - 536
EP - 545
%@ 1673-1581
Y1 - 2008
PB - Zhejiang University Press & Springer
ER -
DOI - 10.1631/jzus.B0720006


Abstract: 
The PhyAm gene encoding acid phytase and optimized neutral phytase PhyCs gene were inserted into expression vector pPIC9K in correct orientation and transformed into Pichia pastoris in order to expand the pH profile of phytase and decrease the cost of production. The fusion phytase PhyAm-PhyCs gene was successfully overexpressed in P. pastoris as an active and extracellular phytase. The yield of total extracellular fusion phytase activity is (25.4±0.53) U/ml at the flask scale and (159.1±2.92) U/ml for high cell-density fermentation, respectively. Purified fusion phytase exhibits an optimal temperature at 55 °C and an optimal pH at 5.5~6.0 and its relative activity remains at a relatively high level of above 70% in the range of pH 2.0 to 7.0. About 51% to 63% of its original activity remains after incubation at 75 °C to 95 °C for 10 min. Due to heavy glycosylation, the expressed fusion phytase shows a broad and diffuse band in SDS-PAGE (sodium dodecyl sulfate-polyacrylamide gel electrophoresis). After deglycosylation by endoglycosidase H (EndoHf), the enzyme has an apparent molecular size of 95 kDa. The characterization of the fusion phytase was compared with those of PhyCs and PhyAm.

Darkslateblue:Affiliate; Royal Blue:Author; Turquoise:Article

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