title: Properties of beta-propeller phytase expressed in transgenic tobacco.
abstract: Phytases are enzymes that liberate inorganic phosphates from phytate. In a previous study, a beta-propeller phytase (168phyA) from Bacillus subtilis was introduced into transgenic tobacco, which resulted in certain phenotypic changes. In the study described herein, the recombinant phytase (t168phyA) was purified from transgenic tobacco to near homogeneity by a three-step purification scheme. The biochemical properties and kinetic parameters of t168phyA were compared with those of its counterpart from B. subtilis. t168phyA was glycosylated, and it showed a 4 kDa increase in molecular size in SDS-PAGE (44 kDa vs. 40 kDa). Although its thermostability remained unchanged, its temperature optimum shifted from 60 degrees C to 45-50 degrees C and its pH optimum shifted from pH 5.5 to 6.0. Kinetic data showed that the t168phyA had a lower Kcat, but a higher Km than the native enzyme. Despite these changes, t168phyA remained catalytically active and has a specific activity of 2.3 U/mg protein. These results verify the activity of recombinant Bacillus phytase that is expressed in plants.
authors.affiliation: Department of Zoology, University of Hong Kong, Pokfulam Road, Hong Kong, China.
authors.forename: Wing-Lee
authors.initials: WL
authors.lastname: Chan
authors.forename: Shiu-Cheung
authors.initials: SC
authors.lastname: Lung
authors.forename: Boon L
authors.initials: BL
authors.lastname: Lim
mesh_terms: D010832:6-Phytase / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D001412:Bacillus subtilis / Q000201:enzymology; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008958:Models, Molecular; D008970:Molecular Weight; D030821:Plants, Genetically Modified; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D013816:Thermodynamics; D014026:Nicotiana / Q000201:enzymology / Q000235:genetics
chemical_list: D001426:Bacterial Proteins; D011994:Recombinant Proteins; D010832:6-Phytase