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Overproduction, purification, crystallization and preliminary X‐ray characterization of the family 46 carbohydrate‐binding module (CBM46) of endo‐β‐1,4‐glucanase B (CelB) from Bacillus halodurans
Author(s) -
Venditto Immacolata,
Santos Helena,
Ferreira Luís M. A.,
Sakka Kazuo,
Fontes Carlos M. G. A.,
Najmudin Shabir
Publication year - 2014
Publication title -
acta crystallographica section f
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.572
H-Index - 37
ISSN - 2053-230X
DOI - 10.1107/s2053230x14008395
Subject(s) - glucanase , escherichia coli , biochemistry , carbohydrate binding module , hemicellulose , cellulose , recombinant dna , protein primary structure , biology , carbohydrate , enzyme , chemistry , microbiology and biotechnology , cellulase , peptide sequence , gene
Plant cell‐wall polysaccharides offer an abundant energy source utilized by many microorganisms, thus playing a central role in carbon recycling. Aerobic microorganisms secrete carbohydrate‐active enzymes (CAZymes) that catabolize this composite structure, comprising cellulose, hemicellulose and lignin, into simple compounds such as glucose. Carbohydrate‐binding modules (CBMs) enhance the efficacy of associated CAZYmes. They are organized into families based on primary‐sequence homology. CBM family 46 contains more than 40 different members, but has yet to be fully characterized. Here, a recombinant derivative of the C‐terminal family 46 CBM module ( Bh CBM46) of Bacillus halodurans endo‐β‐1,4‐glucanase B (CelB) was overexpressed in Escherichia coli and purified by immobilized metal‐ion affinity chromatography. Preliminary structural characterization was carried out on Bh CBM46 crystallized in different conditions. The crystals of Bh CBM46 belonged to the tetragonal space group I 4 1 22. Data were collected for the native form and a selenomethionine derivative to 2.46 and 2.3 Å resolution, respectively. The Bh CBM46 structure was determined by a single‐wavelength anomalous dispersion experiment using AutoSol from the PHENIX suite.