title: Production and characterization of recombinant Phanerochaete chrysosporium beta-glucosidase in the methylotrophic yeast Pichia pastoris.
abstract: The extracellular beta-glucosidase from the white-rot fungus Phanerochaete chrysosporium was expressed heterologously in the methylotrophic yeast Pichia pastoris. After 7 days' cultivation in an induction medium containing 1% (v/v) methanol, the expression level of the recombinant enzyme was 28,500 U/l, 38 times that of the wild-type enzyme. The specific activity of the crude recombinant enzyme for p-nitrophenyl-beta-D-glucoside was 52 U/mg, 37 times that of the wild-type enzyme; this difference made the purification of the enzyme simple. On a SDS-PAGE, the molecular mass of the recombinant enzyme was 133 kDa, and that of the wild-type enzyme was 116 kDa, but the difference had no effect on the hydrolysis of cellobiose or p-nitrophenyl-beta-D-glucoside. We concluded that the recombinant enzyme produced by Pichia pastoris retains the catalytic properties of the wild-type enzyme from Phanerochaete chrysosporium.
authors.affiliation: Graduate School of Agricultural and Life Sciences, University of Tokyo, Yayoi 1-1-1, Bunkyo-ku, Tokyo 113-8657, Japan.
authors.forename: Rie
authors.initials: R
authors.lastname: Kawai
authors.forename: Makoto
authors.initials: M
authors.lastname: Yoshida
authors.forename: Tomomi
authors.initials: T
authors.lastname: Tani
authors.forename: Kiyohiko
authors.initials: K
authors.lastname: Igarashi
authors.forename: Tsuyoshi
authors.initials: T
authors.lastname: Ohira
authors.forename: Hiromichi
authors.initials: H
authors.lastname: Nagasawa
authors.forename: Masahiro
authors.initials: M
authors.lastname: Samejima
mesh_terms: D002482:Cellulose / Q000378:metabolism; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D004591:Electrophoresis, Polyacrylamide Gel; D005822:Genetic Vectors; D007700:Kinetics; D020075:Phanerochaete / Q000201:enzymology* / Q000254:growth & development; D010843:Pichia / Q000201:enzymology*; D011485:Protein Binding; D012331:RNA, Fungal / Q000096:biosynthesis / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis; D001617:beta-Glucosidase / Q000096:biosynthesis*
chemical_list: D012331:RNA, Fungal; D011994:Recombinant Proteins; D002482:Cellulose; D001617:beta-Glucosidase