title: Identification of tandemly repeated type VI cellulose-binding domains in an endoglucanase from the aerobic soil bacterium Cellvibrio mixtus.
abstract: Cellulose-binding domains (CBD) play a pivotal role during plant cell wall hydrolysis by cellulases and xylanases from aerobic soil bacteria. Recently we have reported the molecular characterisation of a single-domain endoglucanase from Cellvibrio mixtus, suggesting that some cellulases produced by this aerobic bacterium preferentially hydrolyse soluble cellulosic substrates. Here we describe the complete nucleotide sequence of a second cellulase gene, celB, from the soil bacterium C. mixtus. It revealed an open reading frame of 1863 bp that encoded a polypeptide, defined as cellulase B (CelB) with a predicted Mr of 66 039. CelB contained a glycosyl hydrolase family 5 catalytic domain at its N terminus followed by two repeated domains, which exhibited sequence identity with type VI CBD previously found in xylanases. Full-length CelB bound to cellulose while catalytically active truncated cellulase derivatives were unable to bind the polysaccharide, confirming that CelB is a modular enzyme and that the type VI CBD homologues were functional. Analysis of the biochemical properties of CelB revealed that the enzyme hydrolyses a range of cellulosic substrates, although it was unable to depolymerise Avicel. We propose that type VI CBD, usually found in xylanases, provide an additional mechanism by which cellulases can accumulate on the surface of the plant cell wall, although they do not potentiate cellulase activity directly. The results demonstrate that C. mixtus, in common with other aerobic bacteria, is able to produce cellulases that are directed to the hydrolysis of cellulose in its natural environment, the plant cell wall.
authors.affiliation: CIISA-Faculdade de Medicina Veterinária, Lisboa, Portugal.
authors.forename: C M
authors.initials: CM
authors.lastname: Fontes
authors.forename: J H
authors.initials: JH
authors.lastname: Clarke
authors.forename: G P
authors.initials: GP
authors.lastname: Hazlewood
authors.forename: T H
authors.initials: TH
authors.lastname: Fernandes
authors.forename: H J
authors.initials: HJ
authors.lastname: Gilbert
authors.forename: L M
authors.initials: LM
authors.lastname: Ferreira
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D001665:Binding Sites; D002480:Cellulase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D002482:Cellulose / Q000378:metabolism*; D016946:Gram-Negative Aerobic Rods and Cocci / Q000201:enzymology*; D008969:Molecular Sequence Data; D012988:Soil Microbiology; D013329:Structure-Activity Relationship
chemical_list: D002482:Cellulose; D002480:Cellulase