Expression and characterization of a GH39 β-xylosidase II from Caulobacter crescentus.

Juliana Moço Corrêa, Luciana Graciano, Josielle Abrahão, Eduardo Alexandre Loth, Rinaldo Ferreira Gandra, Marina Kimiko Kadowaki, Caroline Henn, Rita de Cássia Garcia Simão

Centro de Ciências Exatas e Tecnológicas, Universidade Estadual do Oeste do Paraná, Rua Universitária, 2069, Cascavel, Paraná, 85814-110, Brazil.

Applied biochemistry and biotechnology 2012 Dec

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In the present work, the gene xynB2, encoding a β-xylosidase II of the Glycoside Hydrolase 39 (GH39) family, of Caulobacter crescentus was cloned and successfully overexpressed in Escherichia coli DH10B. The recombinant protein (CcXynB2) was purified using nickel-Sepharose affinity chromatography, with a recovery yield of 75.5 %. CcXynB2 appeared as a single band of 60 kDa on a sodium dodecyl sulfate polyacrylamide gel and was recognized by a specific polyclonal antiserum. The predicted CcXynB2 protein showed a high homology with GH39 β-xylosidases of the genus Xanthomonas. CcXynB2 exhibited an optimal activity at 55 °C and a pH of 6. CcXynB2 displayed stability at pH values of 4.5-7.5 for 24 h and thermotolerance up to 50 °C. The K (M) and V (Max) values were 9.3 ± 0.45 mM and 402 ± 19 μmol min(-1) for ρ-nitrophenyl-β-D-xylopyranoside, respectively. The purified recombinant enzyme efficiently produced reducing sugars from birchwood xylan and sugarcane bagasse fibers pre-treated with a purified xylanase. As few bacterial GH39 family β-xylosidases have been characterized, this work provides a good contribution to this group of enzymes.

Citation

Juliana Moço Corrêa, Luciana Graciano, Josielle Abrahão, Eduardo Alexandre Loth, Rinaldo Ferreira Gandra, Marina Kimiko Kadowaki, Caroline Henn, Rita de Cássia Garcia Simão. Expression and characterization of a GH39 β-xylosidase II from Caulobacter crescentus. Applied biochemistry and biotechnology. 2012 Dec;168(8):2218-29