title: Molecular cloning, overexpression, purification and crystallographic analysis of a GH43 β-xylosidase from Bacillus licheniformis.
abstract: β-Xylosidases (EC 3.2.1.37) catalyze the hydrolysis of short xylooligosaccharides into xylose, which is an essential step in the complete depolymerization of xylan, the major hemicellulosic polysaccharide of plant cell walls, and has great biotechnological relevance for the production of lignocellulose-based biofuels and the paper industry. In this study, a GH43 β-xylosidase identified from the bacterium Bacillus licheniformis (BlXylA) was cloned into the the pET-28a bacterial expression vector, recombinantly overexpressed in Escherichia coli BL21(DE3) cells and purified to homogeneity by metal-affinity and size-exclusion chromatography. The protein was crystallized in the presence of the organic solvent 2-methyl-2,4-pentanediol and a single crystal diffracted to 2.49 Å resolution. The X-ray diffraction data were indexed in the monoclinic space group C2, with unit-cell parameters a = 152.82, b = 41.9, c = 71.79 Å, β = 91.7°. Structural characterization of this enzyme will contribute to a better understanding of the structural requirements for xylooligosaccharide specificity within the GH43 family.
authors.affiliation: Brazilian Bioethanol Science and Technology Laboratory (CTBE), National Center for Research in Energy and Materials (CNPEM), Giuseppe Maximo Scolfaro 10000, 13083-100 Campinas-SP, Brazil.
authors.forename: José Alberto
authors.initials: JA
authors.lastname: Diogo
authors.affiliation: Brazilian Bioethanol Science and Technology Laboratory (CTBE), National Center for Research in Energy and Materials (CNPEM), Giuseppe Maximo Scolfaro 10000, 13083-100 Campinas-SP, Brazil.
authors.forename: Leticia Maria
authors.initials: LM
authors.lastname: Zanphorlin
authors.affiliation: Department of Food Science, University of Campinas, Rua Monteiro Lobato 80, Cidade Universitária, 13081-970 Campinas-SP, Brazil.
authors.forename: Hélia Harumi
authors.initials: HH
authors.lastname: Sato
authors.affiliation: Brazilian Biosciences National Laboratory (LNBio), National Center for Research in Energy and Materials (CNPEM), Giuseppe Maximo Scolfaro 10000, 13083-100 Campinas-SP, Brazil.
authors.forename: Mario Tyago
authors.initials: MT
authors.lastname: Murakami
authors.affiliation: Brazilian Bioethanol Science and Technology Laboratory (CTBE), National Center for Research in Energy and Materials (CNPEM), Giuseppe Maximo Scolfaro 10000, 13083-100 Campinas-SP, Brazil.
authors.forename: Roberto
authors.initials: R
authors.lastname: Ruller
mesh_terms: D000595:Amino Acid Sequence; D001407:Bacillus / Q000737:chemistry* / Q000201:enzymology; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics; D003001:Cloning, Molecular; D003460:Crystallization; D018360:Crystallography, X-Ray; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D006018:Glycols; D008969:Molecular Sequence Data; D011993:Recombinant Fusion Proteins / Q000737:chemistry* / Q000235:genetics; D016415:Sequence Alignment; D012997:Solvents; D014995:Xylosidases / Q000737:chemistry* / Q000235:genetics
chemical_list: D001426:Bacterial Proteins; D006018:Glycols; D011993:Recombinant Fusion Proteins; D012997:Solvents; D014995:Xylosidases; C026579:exo-1,4-beta-D-xylosidase; C011480:hexylene glycol
keywords: Bacillus licheniformis; GH43 family; β-xylosidase