title: Alpha-glucan recognition by a new family of carbohydrate-binding modules found primarily in bacterial pathogens.
abstract: TmPul13, a family 13 glycoside hydrolase from Thermotoga maritima, is a four-module protein having pullulanase activity; the three N-terminal modules are of unknown function while the large C-terminal module is likely the catalytic module. Dissection of the functions of the three unknown modules revealed that the 100 amino acid module at the extreme N-terminus of TmPul13 comprises a new family of carbohydrate-binding modules (CBM) that a bioinformatic analysis shows are most frequently found in pullulanase-like sequences from bacterial pathogens. Detailed binding studies of this isolated CBM, here called TmCBM41, reveals a preference for alpha-(1,4)-linked glucans, but occasional alpha-(1,6)-linked glucose residues, such as those found in pullulan, are tolerated. UV difference, isothermal titration calorimetry, and analytical ultracentrifugation binding studies suggest that maltooligosaccharides longer than four glucose residues are able to bind two TmCBM41 molecules per oligosaccharide when sugar concentrations are below the CBM concentration. This is explained in terms of an equilibrium expression involving the formation of both a 1 to 1 sugar to CBM complex and a 1 to 2 sugar to CBM complex (i.e., a CBM dimer ligated by an oligosaccharide). The presence of an alpha-(1-6) linkage in the oligosaccharide appears to prevent this phenomenon.
authors.affiliation: Department of Biochemistry and Microbiology, University of Victoria, P.O. Box 3055 STN CSC, Victoria, British Columbia V8W 3P6, Canada.
authors.forename: Alicia
authors.initials: A
authors.lastname: Lammerts van Bueren
authors.forename: Ron
authors.initials: R
authors.lastname: Finn
authors.forename: Juan
authors.initials: J
authors.lastname: Ausió
authors.forename: Alisdair B
authors.initials: AB
authors.lastname: Boraston
mesh_terms: D020816:Amino Acid Motifs; D001665:Binding Sites; D002151:Calorimetry; D019295:Computational Biology / Q000379:methods; D005936:Glucans / Q000737:chemistry / Q000378:metabolism*; D006026:Glycoside Hydrolases / Q000737:chemistry / Q000378:metabolism*; D007700:Kinetics; D005810:Multigene Family; D009844:Oligosaccharides / Q000737:chemistry / Q000378:metabolism; D011135:Polysaccharides, Bacterial / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D011956:Receptors, Cell Surface / Q000737:chemistry / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D040681:Structural Homology, Protein; D013816:Thermodynamics; D020124:Thermotoga maritima
chemical_list: D005936:Glucans; D009844:Oligosaccharides; D011135:Polysaccharides, Bacterial; D011956:Receptors, Cell Surface; C021705:maltooligosaccharides; C055213:saccharide-binding proteins; C009109:pullulan; C040088:1,4-glucan; D006026:Glycoside Hydrolases; C100181:pullulanase