Publication: Biochemical characterization and in vitro digestibility assay of Eupenicillium parvum (BCC17694) phytase expressed in Pichia pastoris
Issued Date
2010-03-01
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ISSN
10465928
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2-s2.0-75549086617
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Mahidol University
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SCOPUS
Bibliographic Citation
Protein Expression and Purification. Vol.70, No.1 (2010), 60-67
Suggested Citation
Anusorn Fugthong, Katewadee Boonyapakron, Warasirin Sornlek, Sutipa Tanapongpipat, Lily Eurwilaichitr, Kusol Pootanakit Biochemical characterization and in vitro digestibility assay of Eupenicillium parvum (BCC17694) phytase expressed in Pichia pastoris. Protein Expression and Purification. Vol.70, No.1 (2010), 60-67. doi:10.1016/j.pep.2009.10.001 Retrieved from: https://repository.li.mahidol.ac.th/handle/123456789/28767
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Title
Biochemical characterization and in vitro digestibility assay of Eupenicillium parvum (BCC17694) phytase expressed in Pichia pastoris
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Abstract
A mature phytase cDNA, encoding 441 amino acids, from Eupenicillium parvum (BCC17694) was cloned into a Pichia pastoris expression vector, pPICZαA, and was successfully expressed as active extracellular glycosylated protein. The recombinant phytase contained the active site RHGXRXP and HD sequence motifs, a large α/β domain and a small α-domain that are typical of histidine acid phosphatase. Glycosylation was found to be important for enzyme activity which is most active at 50 °C and pH 5.5. The recombinant phytase displayed broad substrate specificity toward p-nitrophenyl phosphate, sodium-, calcium-, and potassium-phytate. The enzyme lost its activity after incubating at 50 °C for 5 min and is 50% inhibited by 5 mM Cu2+. However, the enzyme exhibits broad pH stability from 2.5 to 8.0 and is resistant to pepsin. In vitro digestibility test suggested that BCC17694 phytase is at least as effective as another recombinant phytase (r-A170) which is comparable to Natuphos, a commercial phytase, in releasing phosphate from corn-based animal feed, suggesting that BCC17694 phytase is suitable for use as phytase supplement in the animal diet. © 2009 Elsevier Inc. All rights reserved.