title: Cloning, expression in Streptomyces lividans and biochemical characterization of a thermostable endo-beta-1,4-xylanase of Thermomonospora alba ULJB1 with cellulose-binding ability.
abstract: Several thermophilic actinomycetes were isolated from urban solid waste. One of them, Thermomonospora alba ULJB1, showed a broad degradative activity on xylan, cellulose, starch and other polymers. Xylanase and cellulase activities were quantified and compared with those Thermomonospora fusca. Genes encoding two different endo-beta-1,4-xylanase were cloned from T. alba ULJB1. One of them, xylA, was sequenced, subcloned and overexpressed in Streptomyces lividans. It encodes a protein of 482 amino acids with a deduced molecular mass of 48,456 Da. The protein contains a 38-amino-acid leader peptide with six Arg+ residues in its amino-terminal end, a catalytic domain and a cellulose-binding domain connected by a linker region rich in proline and glycine. The XylA protein was purified to near homogeneity from S. lividans/XylA cultures. Two forms of the extracellular xylanase, of 48 kDa and 38 kDa, were produced that differed in their cellulose-binding ability. The 48-kDa protein showed a strong binding to cellulose whereas the 38-kDa form did not bind to this polymer, apparently because of the removal during processing of the cellulose-binding domain. Both forms were able to degrade xylans form different origins but not lichenam or carboxymethylcellulose. The major degradation product was xylobiose with traces of xylose. The xylanase activity was thermostable, showing a good activity up to 95 degrees C, and had broad pH stability in the range from pH 4.0 to pH 10.0.
authors.affiliation: Faculty of Biology, University of Léon, Spain. degjmm@unileon.es
authors.forename: J
authors.initials: J
authors.lastname: Blanco
authors.forename: J J
authors.initials: JJ
authors.lastname: Coque
authors.forename: J
authors.initials: J
authors.lastname: Velasco
authors.forename: J F
authors.initials: JF
authors.lastname: Martín
mesh_terms: D000192:Actinomycetales / Q000201:enzymology*; D000595:Amino Acid Sequence; D001483:Base Sequence; D002482:Cellulose / Q000378:metabolism*; D003001:Cloning, Molecular; D043364:Endo-1,4-beta Xylanases; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000378:metabolism; D013302:Streptomyces / Q000235:genetics*; D013379:Substrate Specificity; D014990:Xylans / Q000378:metabolism; D014995:Xylosidases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism
chemical_list: D011994:Recombinant Proteins; D014990:Xylans; D002482:Cellulose; D014995:Xylosidases; D043364:Endo-1,4-beta Xylanases