title: The role of conserved tryptophan residues in the interaction of a bacterial cellulose binding domain with its ligand.
abstract: The five conserved tryptophan residues in the cellulose binding domain of xylanase A from Pseudomonas fluorescens subsp. cellulosa were replaced with alanine and phenylalanine. The mutated domains were fused to mature alkaline phosphatase, and the capacity of the hybrid proteins to bind cellulose was assessed. Alanine substitution of the tryptophan residues, in general, resulted in a significant decrease in the capacity of the cellulose binding domains to bind cellulose. Mutant domains containing phenylalanine substitution retained some affinity for cellulose. The C-terminal proximal tryptophan did not play an important role in ligand binding, while Trp13, Trp34 and Trp38 were essential for the cellulose binding domain to retain cellulose binding capacity. Data presented in this study suggest major differences in the mechanism of cellulose attachment between Pseudomonas and Cellulomonas cellulose binding domains.
authors.affiliation: Department of Biological and Nutritional Sciences, University of Newcastle upon Tyne, UK.
authors.forename: D M
authors.initials: DM
authors.lastname: Poole
authors.forename: G P
authors.initials: GP
authors.lastname: Hazlewood
authors.forename: N S
authors.initials: NS
authors.lastname: Huskisson
authors.forename: R
authors.initials: R
authors.lastname: Virden
authors.forename: H J
authors.initials: HJ
authors.lastname: Gilbert
mesh_terms: D000409:Alanine / Q000494:pharmacology; D000469:Alkaline Phosphatase / Q000096:biosynthesis; D000595:Amino Acid Sequence; D015153:Blotting, Western; D002482:Cellulose / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D043364:Endo-1,4-beta Xylanases; D004926:Escherichia coli; D006026:Glycoside Hydrolases / Q000235:genetics / Q000502:physiology*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010649:Phenylalanine / Q000494:pharmacology; D011551:Pseudomonas fluorescens / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D014364:Tryptophan / Q000502:physiology*
chemical_list: D011993:Recombinant Fusion Proteins; D010649:Phenylalanine; D014364:Tryptophan; D002482:Cellulose; D000469:Alkaline Phosphatase; D006026:Glycoside Hydrolases; D043364:Endo-1,4-beta Xylanases; D000409:Alanine