Open AccessExpression and characterization of Aspergillus thermostable phytases in Pichia pastoris
Author(s) -
Promdonkoy Patcharee,
Tang Kittapong,
Sornlake Warasirin,
Harnpicharnchai Piyanun,
Kobayashi Rutchadaporn Sriprang,
Ruanglek Vasimon,
Upathanpreecha Tewa,
Vesaratchavest Mongkol,
Eurwilaichitr Lily,
Tanapongpipat Sutipa
Publication year - 2009
Publication title -
fems microbiology letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.899
H-Index - 151
eISSN - 1574-6968
pISSN - 0378-1097
DOI - 10.1111/j.1574-6968.2008.01399.x
Subject(s) - phytase , pichia pastoris , thermostability , aspergillus niger , biochemistry , recombinant dna , enzyme , glycosylation , phytic acid , biology , chemistry , food science , gene
Two thermostable phytases were identified from Thai isolates of Aspergillus japonicus BCC18313 (TR86) and Aspergillus niger BCC18081 (TR170). Both genes of 1404 bp length, coding for putative phytases of 468 amino acid residues, were cloned and transferred into Pichia pastoris . The recombinant phytases, r‐PhyA86 and r‐PhyA170, were expressed as active extracellular, glycosylated proteins with activities of 140 and 100 U mL −1 , respectively. Both recombinant phytases exhibited high affinity for phytate but not for p ‐nitrophenyl phosphate. Optimal phytase activity was observed at 50 °C and pH 5.5. High thermostability, which is partly dependent on glycosylation, was demonstrated for both enzymes, as >50% activity was retained after heating at 100 °C for 10 min. The recombinant phytases also exhibited broad pH stability from 2.0 to 8.0 and are resistant to pepsin. In vitro digestibility tests suggested that r‐PhyA86 and r‐PhyA170 are at least as efficient as commercial phytase for hydrolyzing phytate in corn‐based animal feed and are therefore suitable sources of phytase supplement.