The Experts below are selected from a list of 13947 Experts worldwide ranked by ideXlab platform

Bernard Henrissat - One of the best experts on this subject based on the ideXlab platform.

  • a subfamily roadmap of the evolutionarily diverse Glycoside Hydrolase family 16 gh16
    Journal of Biological Chemistry, 2019
    Co-Authors: Alexander Holm Viborg, Nicolas Terrapon, Bernard Henrissat, Vincent Lombard, Gurvan Michel, Mirjam Czjzek
    Abstract:

    Glycoside Hydrolase family (GH) 16 comprises a large and taxonomically diverse family of glycosidases and transglycosidases that adopt a common β-jelly-roll fold and are active on a range of terrestrial and marine polysaccharides. Presently, broadly insightful sequence–function correlations in GH16 are hindered by a lack of a systematic subfamily structure. To fill this gap, we have used a highly scalable protein sequence similarity network analysis to delineate nearly 23,000 GH16 sequences into 23 robust subfamilies, which are strongly supported by hidden Markov model and maximum likelihood molecular phylogenetic analyses. Subsequent evaluation of over 40 experimental three-dimensional structures has highlighted key tertiary structural differences, predominantly manifested in active-site loops, that dictate substrate specificity across the GH16 evolutionary landscape. As for other large GH families (i.e. GH5, GH13, and GH43), this new subfamily classification provides a roadmap for functional glycogenomics that will guide future bioinformatics and experimental structure–function analyses. The GH16 subfamily classification is publicly available in the CAZy database. The sequence similarity network workflow used here, SSNpipe, is freely available from GitHub.

  • substrate specificity regiospecificity and processivity in Glycoside Hydrolase family 74
    Journal of Biological Chemistry, 2019
    Co-Authors: Gregory Arnal, Bernard Henrissat, Alexander Holm Viborg, P J Stogios, Jathavan Asohan, Mohamed A Attia, T Skarina, A Savchenko
    Abstract:

    Glycoside Hydrolase family 74 (GH74) is a historically important family of endo-β-glucanases. On the basis of early reports of detectable activity on cellulose and soluble cellulose derivatives, GH74 was originally considered to be a “cellulase” family, although more recent studies have generally indicated a high specificity toward the ubiquitous plant cell wall matrix glycan xyloglucan. Previous studies have indicated that GH74 xyloglucanases differ in backbone cleavage regiospecificities and can adopt three distinct hydrolytic modes of action: exo, endo-dissociative, and endo-processive. To improve functional predictions within GH74, here we coupled in-depth biochemical characterization of 17 recombinant proteins with structural biology–based investigations in the context of a comprehensive molecular phylogeny, including all previously characterized family members. Elucidation of four new GH74 tertiary structures, as well as one distantly related dual seven-bladed β-propeller protein from a marine bacterium, highlighted key structure–function relationships along protein evolutionary trajectories. We could define five phylogenetic groups, which delineated the mode of action and the regiospecificity of GH74 members. At the extremes, a major group of enzymes diverged to hydrolyze the backbone of xyloglucan nonspecifically with a dissociative mode of action and relaxed backbone regiospecificity. In contrast, a sister group of GH74 enzymes has evolved a large hydrophobic platform comprising 10 subsites, which facilitates processivity. Overall, the findings of our study refine our understanding of catalysis in GH74, providing a framework for future experimentation as well as for bioinformatics predictions of sequences emerging from (meta)genomic studies.

  • How a Glycoside Hydrolase Recognizes a Helical Polyglucan.
    Structure, 2017
    Co-Authors: Bernard Henrissat, Marie-line Garron
    Abstract:

    Similarly to other biopolymers, linear polysaccharides can form double- or triple-helical structures. How enzymes recognize and manage this quaternary structure is an unresolved question. In this issue of Structure , Pluvinage et al. (2017) shed light on the structural complementarity between family GH81 Glycoside Hydrolase and the quaternary structure of their polysaccharide substrates.

  • unusual active site location and catalytic apparatus in a Glycoside Hydrolase family
    Proceedings of the National Academy of Sciences of the United States of America, 2017
    Co-Authors: J Munozmunoz, Alan Cartmell, Nicolas Terrapon, Bernard Henrissat, Harry J. Gilbert
    Abstract:

    The human gut microbiota use complex carbohydrates as major nutrients. The requirement for an efficient glycan degrading systems exerts a major selection pressure on this microbial community. Thus, we propose that these bacteria represent a substantial resource for discovering novel carbohydrate active enzymes. To test this hypothesis, we focused on enzymes that hydrolyze rhamnosidic bonds, as cleavage of these linkages is chemically challenging and there is a paucity of information on l-rhamnosidases. Here we screened the activity of enzymes derived from the human gut microbiota bacterium Bacteroides thetaiotaomicron, which are up-regulated in response to rhamnose-containing glycans. We identified an α-l-rhamnosidase, BT3686, which is the founding member of a Glycoside Hydrolase (GH) family, GH145. In contrast to other rhamnosidases, BT3686 cleaved l-Rha-α1,4–d-GlcA linkages through a retaining double-displacement mechanism. The crystal structure of BT3686 showed that the enzyme displayed a type A seven-bladed β-propeller fold. Mutagenesis and crystallographic studies, including the structure of BT3686 in complex with the reaction product GlcA, revealed a location for the active site among β-propeller enzymes cited on the posterior surface of the rhamnosidase. In contrast to the vast majority of GH, the catalytic apparatus of BT3686 does not comprise a pair of carboxylic acid residues but, uniquely, a single histidine functions as the only discernable catalytic amino acid. Intriguingly, the histidine, His48, is not invariant in GH145; however, when engineered into structural homologs lacking the imidazole residue, α-l-rhamnosidase activity was established. The potential contribution of His48 to the catalytic activity of BT3686 is discussed.

  • Unusual active site location and catalytic apparatus in a Glycoside Hydrolase family
    Proceedings of the National Academy of Sciences of the United States of America, 2017
    Co-Authors: Jose Munoz-munoz, Alan Cartmell, Nicolas Terrapon, Bernard Henrissat, Harry J. Gilbert
    Abstract:

    The human gut microbiota use complex carbohydrates as major nutrients. The requirement for an efficient glycan degrading systems exerts a major selection pressure on this microbial community. Thus, we propose that these bacteria represent a substantial resource for discovering novel carbohydrate active enzymes. To test this hypothesis, we focused on enzymes that hydrolyze rhamnosidic bonds, as cleavage of these linkages is chemically challenging and there is a paucity of information on L-rhamnosidases. Here we screened the activity of enzymes derived from the human gut microbiota bacterium Bacteroides thetaiotaomicron, which are up-regulated in response to rhamnose-containing glycans. We identified an alpha-L-rhamnosidase, BT3686, which is the founding member of a Glycoside Hydrolase (GH) family, GH145. In contrast to other rhamnosidases, BT3686 cleaved L-Rh alpha-a1,4-D-GlcA linkages through a retaining double-displacement mechanism. The crystal structure of BT3686 showed that the enzyme displayed a type A sevenbladed beta-propeller fold. Mutagenesis and crystallographic studies, including the structure of BT3686 in complex with the reaction product GlcA, revealed a location for the active site among beta-propeller enzymes cited on the posterior surface of the rhamnosidase. In contrast to the vast majority of GH, the catalytic apparatus of BT3686 does not comprise a pair of carboxylic acid residues but, uniquely, a single histidine functions as the only discernable catalytic amino acid. Intriguingly, the histidine, His48, is not invariant in GH145; however, when engineered into structural homologs lacking the imidazole residue, alpha-L-rhamnosidase activity was established. The potential contribution of His48 to the catalytic activity of BT3686 is discussed.

Jan-hendrik Hehemann - One of the best experts on this subject based on the ideXlab platform.

Craig S. Robb - One of the best experts on this subject based on the ideXlab platform.

Zui Fujimoto - One of the best experts on this subject based on the ideXlab platform.

  • crystal structure and characterization of the Glycoside Hydrolase family 62 α l arabinofuranosidase from streptomyces coelicolor
    Journal of Biological Chemistry, 2014
    Co-Authors: Tomoko Maehara, Zui Fujimoto, Hitomi Ichinose, M Michikawa, Koichi Harazono, Satoshi Kaneko
    Abstract:

    α-l-Arabinofuranosidase, which belongs to the Glycoside Hydrolase family 62 (GH62), hydrolyzes arabinoxylan but not arabinan or arabinogalactan. The crystal structures of several α-l-arabinofuranosidases have been determined, although the structures, catalytic mechanisms, and substrate specificities of GH62 enzymes remain unclear. To evaluate the substrate specificity of a GH62 enzyme, we determined the crystal structure of α-l-arabinofuranosidase, which comprises a carbohydrate-binding module family 13 domain at its N terminus and a catalytic domain at its C terminus, from Streptomyces coelicolor. The catalytic domain was a five-bladed β-propeller consisting of five radially oriented anti-parallel β-sheets. Sugar complex structures with l-arabinose, xylotriose, and xylohexaose revealed five subsites in the catalytic cleft and an l-arabinose-binding pocket at the bottom of the cleft. The entire structure of this GH62 family enzyme was very similar to that of Glycoside Hydrolase 43 family enzymes, and the catalytically important acidic residues found in family 43 enzymes were conserved in GH62. Mutagenesis studies revealed that Asp202 and Glu361 were catalytic residues, and Trp270, Tyr461, and Asn462 were involved in the substrate-binding site for discriminating the substrate structures. In particular, hydrogen bonding between Asn462 and xylose at the nonreducing end subsite +2 was important for the higher activity of substituted arabinofuranosyl residues than that for terminal arabinofuranoses.

  • Bacteroides thetaiotaomicron VPI‐5482 Glycoside Hydrolase family 66 homolog catalyzes dextranolytic and cyclization reactions
    FEBS Journal, 2012
    Co-Authors: Eiji Yamamoto, Hiroyuki Nakai, Min Sun Kang, Wataru Saburi, Masayuki Okuyama, Haruhide Mori, Kazumi Funane, Mitsuru Momma, Zui Fujimoto
    Abstract:

    Bacteroides thetaiotaomicron VPI-5482 harbors a gene encoding a putative cycloisomaltooligosaccharide glucanotransferase (BT3087) belonging to Glycoside Hydrolase family 66. The goal of the present study was to characterize the catalytic properties of this enzyme. Therefore, we expressed BT3087 (recombinant endo-dextranase from Bacteroides thetaiotaomicron VPI-5482) in Escherichia coli and determined that recombinant endo-dextranase from Bacteroides thetaiotaomicron VPI-5482 preferentially synthesized isomaltotetraose and isomaltooligosaccharides (degree of polymerization > 4) from dextran. The enzyme also generated large cyclic isomaltooligosaccharides early in the reaction. We conclude that members of the Glycoside Hydrolase 66 family may be classified into three types: (a) endo-dextranases, (b) dextranases possessing weak cycloisomaltooligosaccharide glucanotransferase activity, and (c) cycloisomaltooligosaccharide glucanotransferases.

  • structural elucidation of dextran degradation mechanism by streptococcus mutans dextranase belonging to Glycoside Hydrolase family 66
    Journal of Biological Chemistry, 2012
    Co-Authors: Nobuhiro Suzuki, Masayuki Okuyama, Haruhide Mori, Kazumi Funane, Mitsuru Momma, Zui Fujimoto, Atsuo Kimura
    Abstract:

    Abstract Dextranase is an enzyme that hydrolyzes dextran α-1,6 linkages. Streptococcus mutans dextranase (SmDex) belongs to Glycoside Hydrolase family 66, producing isomaltooligosaccharides of various sizes, and consisting of at least five amino acid sequence regions. The crystal structure of the conserved fragment from Gln-100 to Ile-732 of SmDex, devoid of its N and C-terminal variable regions, was determined at 1.6 A resolution and found to contain three structural domains. Domain N possessed an immunoglobulin-like β-sandwich fold, domain A the enzyme's catalytic module, comprising a (β/α)8-barrel, and domain C formed a β-sandwich structure containing two Greek key motifs. Two ligand complex structures were also determined and, in the enzyme/isomaltotriose complex structure, the bound isomaltooligosaccharide with four glucose moieties was observed in the catalytic glycone cleft and considered to be the transglycosylation product of the enzyme, indicating the presence of four subsites -4 to -1 in the catalytic cleft. The complexed structure with 4′,5′-epoxypentyl-α-D-glucopyranoside, a suicide substrate of the enzyme, revealed that the epoxide ring reacted to form a covalent bond with the Asp-385 sidechain. These structures collectively indicated that Asp-385 was the catalytic nucleophile and Glu-453 the acid/base of the double displacement mechanism, in which the enzyme showed a retaining catalytic character. This is the first structural report for the enzyme belonging to Glycoside Hydrolase family 66, elucidating the enzyme's catalytic machinery.

  • structural and biochemical characterization of Glycoside Hydrolase family 79 β glucuronidase from acidobacterium capsulatum
    Journal of Biological Chemistry, 2012
    Co-Authors: M Michikawa, Mitsuru Momma, Hitomi Ichinose, Stephen G Withers, Peter Biely, Seino A K Jongkees, Makoto Yoshida, Toshihisa Kotake, Yoichi Tsumuraya, Zui Fujimoto
    Abstract:

    Abstract We present the first structure of a Glycoside Hydrolase family 79 β-glucuronidase from Acidobacterium capsulatum, both as a product complex with β-d-glucuronic acid (GlcA) and as its trapped covalent 2-fluoroglucuronyl intermediate. This enzyme consists of a catalytic (β/α)8-barrel domain and a β-domain with irregular Greek key motifs that is of unknown function. The enzyme showed β-glucuronidase activity and trace levels of β-glucosidase and β-xylosidase activities. In conjunction with mutagenesis studies, these structures identify the catalytic residues as Glu173 (acid base) and Glu287 (nucleophile), consistent with the retaining mechanism demonstrated by 1H NMR analysis. Glu45, Tyr243, Tyr292–Gly294, and Tyr334 form the catalytic pocket and provide substrate discrimination. Consistent with this, the Y292A mutation, which affects the interaction between the main chains of Gln293 and Gly294 and the GlcA carboxyl group, resulted in significant loss of β-glucuronidase activity while retaining the side activities at wild-type levels. Likewise, although the β-glucuronidase activity of the Y334F mutant is ∼200-fold lower (kcat/Km) than that of the wild-type enzyme, the β-glucosidase activity is actually 3 times higher and the β-xylosidase activity is only 2.5-fold lower than the equivalent parameters for wild type, consistent with a role for Tyr334 in recognition of the C6 position of GlcA. The involvement of Glu45 in discriminating against binding of the O-methyl group at the C4 position of GlcA is revealed in the fact that the E45D mutant hydrolyzes PNP-β-GlcA approximately 300-fold slower (kcat/Km) than does the wild-type enzyme, whereas 4-O-methyl-GlcA-containing oligosaccharides are hydrolyzed only 7-fold slower.

  • crystal structure of an exo 1 5 α l arabinofuranosidase from streptomyces avermitilis provides insights into the mechanism of substrate discrimination between exo and endo type enzymes in Glycoside Hydrolase family 43
    Journal of Biological Chemistry, 2010
    Co-Authors: Zui Fujimoto, Motomitsu Kitaoka, Tomoko Maehara, Hitomi Ichinose, Mariko Honda, Satoshi Kaneko
    Abstract:

    Exo-1,5-α-l-arabinofuranosidases belonging to Glycoside Hydrolase family 43 have strict substrate specificity. These enzymes hydrolyze only the α-1,5-linkages of linear arabinan and arabino-oligosaccharides in an exo-acting manner. The enzyme from Streptomyces avermitilis contains a core catalytic domain belonging to Glycoside Hydrolase family 43 and a C-terminal arabinan binding module belonging to carbohydrate binding module family 42. We determined the crystal structure of intact exo-1,5-α-l-arabinofuranosidase. The catalytic module is composed of a 5-bladed β-propeller topologically identical to the other family 43 enzymes. The arabinan binding module had three similar subdomains assembled against one another around a pseudo-3-fold axis, forming a β-trefoil-fold. A sugar complex structure with α-1,5-l-arabinofuranotriose revealed three subsites in the catalytic domain, and a sugar complex structure with α-l-arabinofuranosyl azide revealed three arabinose-binding sites in the carbohydrate binding module. A mutagenesis study revealed that substrate specificity was regulated by residues Asn-159, Tyr-192, and Leu-289 located at the aglycon side of the substrate-binding pocket. The exo-acting manner of the enzyme was attributed to the strict pocket structure of subsite −1, formed by the flexible loop region Tyr-281–Arg-294 and the side chain of Tyr-40, which occupied the positions corresponding to the catalytic glycon cleft of GH43 endo-acting enzymes.

Harry Brumer - One of the best experts on this subject based on the ideXlab platform.

  • growth of chitinophaga pinensis on plant cell wall glycans and characterisation of a Glycoside Hydrolase family 27 β l arabinopyranosidase implicated in arabinogalactan utilisation
    PLOS ONE, 2015
    Co-Authors: Lauren S Mckee, Harry Brumer
    Abstract:

    The genome of the soil bacterium Chitinophaga pinensis encodes a diverse array of carbohydrate active enzymes, including nearly 200 representatives from over 50 Glycoside Hydrolase (GH) families, the enzymology of which is essentially unexplored. In light of this genetic potential, we reveal that C. pinensis has a broader saprophytic capacity to thrive on plant cell wall polysaccharides than previously reported, and specifically that secretion of β-l-arabinopyranosidase activity is induced during growth on arabinogalactan. We subsequently correlated this activity with the product of the Cpin_5740 gene, which encodes the sole member of Glycoside Hydrolase family 27 (GH27) in C. pinensis, CpArap27. Historically, GH27 is most commonly associated with α-d-galactopyranosidase and α-d-N-acetylgalactosaminidase activity. A new phylogenetic analysis of GH27 highlighted the likely importance of several conserved secondary structural features in determining substrate specificity and provides a predictive framework for identifying enzymes with the less common β-l-arabinopyranosidase activity.

  • Enzymatic characterization of a Glycoside Hydrolase family 5 subfamily 7 (GH5_7) mannanase from Arabidopsis thaliana
    Planta, 2013
    Co-Authors: Yang Wang, Francisco Vilaplana, Harry Brumer, Henrik Aspeborg
    Abstract:

    Each plant genome contains a repertoire of β-mannanase genes belonging to Glycoside Hydrolase family 5 subfamily 7 (GH5_7), putatively involved in the degradation and modification of various plant mannan polysaccharides, but very few have been characterized at the gene product level. The current study presents recombinant production and in vitro characterization of AtMan5-1 as a first step towards the exploration of the catalytic capacity of Arabidopsis thaliana β-mannanase. The target enzyme was expressed in both E. coli (AtMan5-1e) and P. pastoris (AtMan5-1p). The main difference between the two forms was a higher observed thermal stability for AtMan5-1p, presumably due to glycosylation of that particular variant. AtMan5-1 displayed optimal activity at pH 5 and 35 °C and hydrolyzed polymeric carob galactomannan, konjac glucomannan, and spruce galactoglucomannan as well as oligomeric mannopentaose and mannohexaose. However, the galactose-rich and highly branched guar gum was not as efficiently degraded. AtMan5-1 activity was enhanced by Co(2+) and inhibited by Mn(2+). The catalytic efficiency values for carob galactomannan were 426.8 and 368.1 min(-1) mg(-1) mL for AtMan5-1e and AtMan5-1p, respectively. Product analysis of AtMan5-1p suggested that at least five substrate-binding sites were required for manno-oligosaccharide hydrolysis, and that the enzyme also can act as a transglycosylase.

  • structural enzymology of cellvibrio japonicus agd31b protein reveals α transglucosylase activity in Glycoside Hydrolase family 31
    Journal of Biological Chemistry, 2012
    Co-Authors: Johan Larsbrink, Harry Brumer, A Izumi, G R Hemsworth, G J Davies
    Abstract:

    Abstract The metabolism of the storage polysaccharides glycogen and starch are of vital importance to organisms from all domains of life. In bacteria, utilization of these α-glucans requires the concerted action of a variety of enzymes, including Glycoside Hydrolases, Glycoside phosphorylases, and transglycosylases. In particular, transglycosylases from Glycoside Hydrolase families GH13 and GH77 play well-established roles in α-glucan sidechain (de)branching, regulation of oligo- and polysaccharide chain length, and formation of cyclic dextrans. Here, we present the biochemical and tertiary structural characterization of a new type of bacterial 1,4-α-glucan 4-α-glucosyltransferase from Glycoside Hydrolase family 31 (GH31). Distinct from 1,4-α-glucan 6-α-glucosyltransferases (EC 2.4.1.24) and 4-α-glucanotransferases (EC 2.4.1.25), this enzyme strictly transferred one glucosyl residue from α(1 →4) glucans in disproportionation reactions. Substrate hydrolysis was undetectable for a series of malto-oligosaccharides, except maltose, for which transglycosylation nonetheless dominated across a range of substrate concentrations. Crystallographic analysis of the enzyme in apo, acarbose-complexed, and trapped 5-β-fluoroglucosyl-enzyme intermediate forms revealed extended substrate interactions across one negative and up to three positive subsites, thus providing structural rationalization for the unique, single-monosaccharide transferase activity of the enzyme.

  • evolution substrate specificity and subfamily classification of Glycoside Hydrolase family 5 gh5
    BMC Evolutionary Biology, 2012
    Co-Authors: Henrik Aspeborg, Harry Brumer, Pedro M. Coutinho, Yang Wang, Bernard Henrissat
    Abstract:

    Background The large Glycoside Hydrolase family 5 (GH5) groups together a wide range of enzymes acting on β-linked oligo- and polysaccharides, and glycoconjugates from a large spectrum of organisms. The long and complex evolution of this family of enzymes and its broad sequence diversity limits functional prediction. With the objective of improving the differentiation of enzyme specificities in a knowledge-based context, and to obtain new evolutionary insights, we present here a new, robust subfamily classification of family GH5.