title: Heterologous expression of β-xylosidase gene from Paecilomyces thermophila in Pichia pastoris.
abstract: β-xylosidase from thermophilic fungi Paecilomyces thermophila was functionally expressed in Pichia pastoris with a his tag in the C-terminal under the alcohol oxidase 1 (AOX1) promoter and secreted into the medium at 0.22 mg l(-1). Its molecular mass was estimated to be 52.3 kDa based on the SDS-PAGE analysis, which is 1.3 times higher than the predicted 39.31 kDa from its amino acid compositions, although no potential N- or O- glycosylation sites were predicted from its amino acid sequence. This is presumed to be caused by some unpredictable posttranslational modifications based on mass spectrum analysis of the recombinant protein. The enzyme was most active at 60 °C and pH 7. It showed not only a β-xylosidase activity with a K(m) of 8 mM and a V(max) of 54 μmol min(-1) mg(-1) for hydrolysis of p-nitrophenyl β-D-xylopyranoside but also an arabinofuranosidase activity (6.2 U mg(-1)) on p-nitrophenyl arabinofuranoside.
authors.affiliation: Institute of Chemical and Engineering Sciences, Agency for Science, Technology and Research (A STAR), 1 Pesek Road, Jurong Island, Singapore 627833, Singapore.
authors.forename: Veeresh
authors.initials: V
authors.lastname: Juturu
authors.forename: Jin Chuan
authors.initials: JC
authors.lastname: Wu
mesh_terms: D000595:Amino Acid Sequence; D003001:Cloning, Molecular; D004795:Enzyme Stability; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D015870:Gene Expression; D007700:Kinetics; D008969:Molecular Sequence Data; D010143:Paecilomyces / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics; D010843:Pichia / Q000235:genetics* / Q000378:metabolism; D016415:Sequence Alignment; D013379:Substrate Specificity; D014995:Xylosidases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism
chemical_list: D005656:Fungal Proteins; D014995:Xylosidases