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B Henrissat

Publications and source records attributed to B Henrissat.

At least 19 recordsLinked to original sources
PubMed2006

The genome of black cottonwood, Populus trichocarpa (Torr. & Gray).

We report the draft genome of the black cottonwood tree, Populus trichocarpa. Integration of shotgun sequence assembly with genetic mapping enabled chromosome-scale reconstruction of the genome. More than 45,000 putative protein-coding genes were identified. Analysis of the assembled genome revealed a whole-genome duplication event; about 8000 pairs of duplicated genes from that event survived in the Populus genome. A second, older duplication event is indistinguishably coincident with the divergence of the Populus and Arabidopsis lineages. Nucleotide substitution, tandem gene duplication, and gross chromosomal rearrangement appear to proceed substantially more slowly in Populus than in Arabidopsis. Populus has more protein-coding genes than Arabidopsis, ranging on average from 1.4 to 1.6 putative Populus homologs for each Arabidopsis gene. However, the relative frequency of protein domains in the two genomes is similar. Overrepresented exceptions in Populus include genes associated with lignocellulosic wall biosynthesis, meristem development, disease resistance, and metabolite transport.

Arabidopsis↗
PubMed2002

Complexities in ETS-domain transcription factor function and regulation: lessons from the TCF (ternary complex factor) subfamily. The Colworth Medal Lecture.

The ETS-domain transcription factor family can be divided into a series of subfamilies. Elk-1 represents the founding member of the ternary complex factor (TCF) subfamily. By focusing on the TCF subfamily, we can demonstrate the complexities that exist in the function and regulation of ETS-domain transcription factors. This article focuses on Elk-1 in detail and summarizes the functions of other TCFs. The key themes covered include the domain structure of the TCFs, the mechanisms of complex formation with serum response factor, regulation of TCFs by mitogen-activated protein kinase cascades, and transcriptional regulatory properties of the TCFs. Finally, the emerging role of the TCFs in vivo is discussed. A picture is developing indicating that, while these proteins exhibit significant sequence and functional conservation, key differences in their structure and regulation are being identified which may relate to unique functions of these proteins in vivo.

Amino Acid Sequence↗
PubMed2001

The location of the ligand-binding site of carbohydrate-binding modules that have evolved from a common sequence is not conserved.

Polysaccharide-degrading enzymes are generally modular proteins that contain non-catalytic carbohydrate-binding modules (CBMs), which potentiate the activity of the catalytic module. CBMs have been grouped into sequence-based families, and three-dimensional structural data are available for half of these families. Clostridium thermocellum xylanase 11A is a modular enzyme that contains a CBM from family 6 (CBM6), for which no structural data are available. We have determined the crystal structure of this module to a resolution of 2.1 A. The protein is a beta-sandwich that contains two potential ligand-binding clefts designated cleft A and B. The CBM interacts primarily with xylan, and NMR spectroscopy coupled with site-directed mutagenesis identified cleft A, containing Trp-92, Tyr-34, and Asn-120, as the ligand-binding site. The overall fold of CBM6 is similar to proteins in CBM families 4 and 22, although surprisingly the ligand-binding site in CBM4 and CBM22 is equivalent to cleft B in CBM6. These structural data define a superfamily of CBMs, comprising CBM4, CBM6, and CBM22, and demonstrate that, although CBMs have evolved from a relatively small number of ancestors, the structural elements involved in ligand recognition have been assembled at different locations on the ancestral scaffold.

Amino Acid Sequence↗
PubMed2001

The iota-carrageenase of Alteromonas fortis. A beta-helix fold-containing enzyme for the degradation of a highly polyanionic polysaccharide.

Carrageenans are gel-forming hydrocolloids extracted from the cell walls of marine red algae. They consist of d-galactose residues bound by alternate alpha(1-->3) and beta(1-->4) linkages and substituted by one (kappa-carrageenan), two (iota-carrageenan), or three (lambda-carrageenan) sulfate-ester groups per disaccharide repeating unit. Both the kappa- and iota-carrageenan chains adopt ordered conformations leading to the formation of highly ordered aggregates of double-stranded helices. Several kappa-carrageenases and iota-carrageenases have been cloned from marine bacteria. Kappa-carrageenases belong to family 16 of the glycoside hydrolases, which essentially encompasses polysaccharidases specialized in the hydrolysis of the neutral polysaccharides such as agarose, laminarin, lichenan, and xyloglucan. In contrast, iota-carrageenases constitute a novel glycoside hydrolase structural family. We report here the crystal structure of Alteromonas fortis iota-carrageenase at 1.6 A resolution. The enzyme folds into a right-handed parallel beta-helix of 10 complete turns with two additional C-terminal domains. Glu(245), Asp(247), or Glu(310), in the cleft of the enzyme, are proposed as candidate catalytic residues. The protein contains one sodium and one chloride binding site and three calcium binding sites shown to be involved in stabilizing the enzyme structure.

Alteromonas↗
PubMed2001

Enzyme-catalyzed condensation reaction in a mammalian alpha-amylase. High-resolution structural analysis of an enzyme-inhibitor complex.

Mammalian alpha-amylases catalyze the hydrolysis of alpha-linked glucose polymers according to a complex processive mechanism. We have determined the X-ray structures of porcine pancreatic alpha-amylase complexes with the smallest molecule of the trestatin family (acarviosine-glucose) which inhibits porcine pancreatic alpha-amylase and yet is not hydrolyzed by the enzyme. A structure analysis at 1.38 A resolution of this complex allowed for a clear identification of a genuine single hexasaccharide species composed of two alpha-1,4-linked original molecules bound to the active site of the enzyme. The structural results supported by mass spectrometry experiments provide evidence for an enzymatically catalyzed condensation reaction in the crystal.

Acarbose↗
PubMed2001

Streptomyces matensis laminaripentaose hydrolase is an 'inverting' beta-1,3-glucanase.

The laminaripentaose-producing beta-1,3-glucanase of Streptomyces matensis is a member of the glycoside hydrolase family GH-64. We have constructed and purified a recombinant hexahistidine-tagged form of the enzyme for characterisation. The enzyme, which exists as a monomer in solution, hydrolyses beta-1,3-glucan by a mechanism leading to overall inversion of the anomeric configuration. This is the first determination of the mechanism prevailing in glycoside hydrolase family GH-64 and this is the first characterisation of an 'inverting' beta-1,3-glucanase.

Biopolymers↗
PubMed2001

Crystal structure of a monocotyledon (maize ZMGlu1) beta-glucosidase and a model of its complex with p-nitrophenyl beta-D-thioglucoside.

The maize beta-glucosidase isoenzymes ZMGlu1 and ZMGlu2 hydrolyse the abundant natural substrate DIMBOAGlc (2-O-beta-D-glucopyranosyl-4-hydroxy-7-methoxy-1,4-benzoxazin-3-one), whose aglycone DIMBOA (2,4-hydroxy-7-methoxy-1,4-benzoxazin-3-one) is the major defence chemical protecting seedlings and young plant parts against herbivores and other pests. The two isoenzymes hydrolyse DIMBOAGlc with similar kinetics but differ from each other and their sorghum homologues with respect to specificity towards other substrates. To gain insights into the mechanism of substrate (i.e. aglycone) specificity between the two maize isoenzymes and their sorghum homologues, ZMGlu1 was produced in Escherichia coli, purified, crystallized and its structure solved at 2.5 Angstrom resolution by X-ray crystallography. In addition, the complex of ZMGlu1 with the non-hydrolysable inhibitor p-nitrophenyl beta-D-thioglucoside was crystallized and, based on the partial electron density, a model for the inhibitor molecule within the active site is proposed. The inhibitor is located in a slot-like active site where its aromatic aglycone is held by stacking interactions with Trp-378. Whereas some of the atoms on the non-reducing end of the glucose moiety can be modelled on the basis of the electron density, most of the inhibitor atoms are highly disordered. This is attributed to the requirement of the enzyme to accommodate two different species, namely the substrate in its ground state and in its distorted conformation, for catalysis.

Amino Acid Sequence↗
PubMed2001

Optimized mixtures of recombinant Humicola insolens cellulases for the biodegradation of crystalline cellulose.

The digestion of bacterial cellulose ribbons by ternary mixtures of enzymes consisting of recombinant cellulases (two cellobiohydrolases, Cel6A and Cel7A, and the endoglucanase Cel45A) from Humicola insolens was investigated over a wide range of mixture composition. The extent of digestion was followed by soluble sugar release (saccharification) analysis together with transmission electron microscopy (TEM) observations. It was found that the addition of minute quantities of Cel45A induced a spectacular increase in saccharification of the substrate with either Cel7A or the mixture of Cel6A and Cel7A. Conversely, only a moderate saccharification resulted from the mixing of Cel45A and Cel6A. This difference is believed to originate from (1) the occasional endo character of Cel6A and (2) the competition of Cel6A and Cel45A for the substrate sites that are sensitive to endo activity. Interestingly, the mixture of enzymes giving rise to the highest saccharification rate did not always correspond to mixtures of enzymes generating the highest synergy. TEM images revealed that the bacterial cellulose ribbons became at the same time cut and narrowed down under the action of an optimized mixture of the three enzymes.

Ascomycota↗
PubMed2001

Glycoside hydrolases and glycosyltransferases: families and functional modules.

The past year has witnessed the expected increase in the number of solved structures of glycoside hydrolases and glycosyltransferases, and their constitutive modules. These structures show that, while glycoside hydrolases display an extraordinary variety of folds, glycosyltransferases and carbohydrate-binding modules appear to belong to a much smaller number of folding families.

Animals↗
PubMed2001

The kappa-carrageenase of P. carrageenovora features a tunnel-shaped active site: a novel insight in the evolution of Clan-B glycoside hydrolases.

BACKGROUND: kappa-carrageenans are gel-forming, sulfated 1,3-alpha-1,4-beta-galactans from the cell walls of marine red algae. The kappa-carrageenase from the marine, gram-negative bacterium Pseudoalteromonas carrageenovora degrades kappa-carrageenan both in solution and in solid state by an endoprocessive mechanism. This beta-galactanase belongs to the clan-B of glycoside hydrolases. RESULTS: The structure of P. carrageenovora kappa-carrageenase has been solved to 1.54 A resolution by the multiwavelength anomalous diffraction (MAD) method, using a seleno-methionine-substituted form of the enzyme. The enzyme folds into a curved beta sandwich, with a tunnel-like active site cavity. Another remarkable characteristic is the presence of an arginine residue at subsite -1. CONCLUSIONS: The crystal structure of P. carrageenovora kappa-carrageenase is the first three-dimensional structure of a carrageenase. Its tunnel-shaped active site, the first to be reported for enzymes other than cellulases, suggests that such tunnels are associated with the degradation of solid polysaccharides. Clan-B glycoside hydrolases fall into two subgroups, one with catalytic machinery held by an ancestral beta bulge, and the other in which it is held by a regular beta strand. At subsite -1, all of these hydrolases exhibit an aromatic amino acid that interacts with the hexopyranose ring of the monosaccharide undergoing catalysis. In addition, in kappa-carrageenases, an arginine residue recognizes the sulfate-ester substituents of the beta-linked kappa-carrageenan monomers. It also appears that, in addition to the nucleophile and acid/base catalysts, two other amino acids are involved with the catalytic cycle, accelerating the deglycosylation step.

Alteromonas↗
PubMed2001

Glycosidase active site mutations in human alpha-L-iduronidase.

Mucopolysaccharidosis type I (MPS I; McKusick 25280) results from a deficiency in alpha-L-iduronidase activity. Using a bioinformatics approach, we have previously predicted the putative acid/base catalyst and nucleophile residues in the active site of this human lysosomal glycosidase to be Glu182 and Glu299, respectively. To obtain experimental evidence supporting these predictions, wild-type alpha-L-iduronidase and site-directed mutants E182A and E299A were individually expressed in Chinese hamster ovary-K1 cell lines. We have compared the synthesis, processing, and catalytic properties of the two mutant proteins with wild-type human alpha-L-iduronidase. Both E182A and E299A transfected cells produced catalytically inactive human alpha-L-iduronidase protein at levels comparable to the wild-type control. The E182A protein was synthesized, processed, targeted to the lysosome, and secreted in a similar fashion to wild-type alpha-L-iduronidase. The E299A mutant protein was also synthesized and secreted similarly to the wild-type enzyme, but there were alterations in its rate of traffic and proteolytic processing. These data indicate that the enzymatic inactivity of the E182A and E299A mutants is not due to problems of synthesis/folding, but to the removal of key catalytic residues. In addition, we have identified a MPS I patient with an E182K mutant allele. The E182K mutant protein was expressed in CHO-K1 cells and also found to be enzymatically inactive. Together, these results support the predicted role of E182 and E299 in the catalytic mechanism of alpha-L-iduronidase and we propose that the mutation of either of these residues would contribute to a very severe clinical phenotype in a MPS I patient.

Amino Acid Sequence↗
PubMed2001

The celA gene, encoding a glycosyl hydrolase family 3 beta-glucosidase in Azospirillum irakense, is required for optimal growth on cellobiosides.

The CelA beta-glucosidase of Azospirillum irakense, belonging to glycosyl hydrolase family 3 (GHF3), preferentially hydrolyzes cellobiose and releases glucose units from the C(3), C(4), and C(5) oligosaccharides. The growth of a DeltacelA mutant on these cellobiosides was affected. In A. irakense, the GHF3 beta-glucosidases appear to be functional alternatives for the GHF1 beta-glucosidases in the assimilation of beta-glucosides by other bacteria.

Amino Acid Sequence↗
PubMed2001

A census of carbohydrate-active enzymes in the genome of Arabidopsis thaliana.

The synthesis, modification, and breakdown of carbohydrates is one of the most fundamentally important reactions in nature. The structural and functional diversity of glycosides is mirrored by a vast array of enzymes involved in their synthesis (glycosyltransferases), modification (carbohydrate esterases) and breakdown (glycoside hydrolases and polysaccharide lyases). The importance of these processes is reflected in the dedication of 1-2% of an organism's genes to glycoside hydrolases and glycosyltransferases alone. In plants, these processes are of particular importance for cell-wall synthesis and expansion. starch metabolism, defence against pathogens, symbiosis and signalling. Here we present an analysis of over 730 open reading frames representing the two main classes of carbohydrate-active enzymes, glycoside hydrolases and glycosyltransferases, in the genome of Arabidopsis thaliana. The vast importance of these enzymes in cell-wall formation and degradation is revealed along with the unexpected dominance of pectin degradation in Arabidopsis, with at least 170 open-reading frames dedicated solely to this task.

Arabidopsis↗
PubMed2000

High resolution X-ray crystallography shows that ascorbate is a cofactor for myrosinase and substitutes for the function of the catalytic base.

Myrosinase, an S-glycosidase, hydrolyzes plant anionic 1-thio-beta-d-glucosides (glucosinolates) considered part of the plant defense system. Although O-glycosidases are ubiquitous, myrosinase is the only known S-glycosidase. Its active site is very similar to that of retaining O-glycosidases, but one of the catalytic residues in O-glycosidases, a carboxylate residue functioning as the general base, is replaced by a glutamine residue. Myrosinase is strongly activated by ascorbic acid. Several binary and ternary complexes of myrosinase with different transition state analogues and ascorbic acid have been analyzed at high resolution by x-ray crystallography along with a 2-deoxy-2-fluoro-glucosyl enzyme intermediate. One of the inhibitors, d-gluconhydroximo-1,5-lactam, binds simultaneously with a sulfate ion to form a mimic of the enzyme-substrate complex. Ascorbate binds to a site distinct from the glucose binding site but overlapping with the aglycon binding site, suggesting that activation occurs at the second step of catalysis, i.e. hydrolysis of the glycosyl enzyme. A water molecule is placed perfectly for activation by ascorbate and for nucleophilic attack on the covalently trapped 2-fluoro-glucosyl-moiety. Activation of the hydrolysis of the glucosyl enzyme intermediate is further evidenced by the observation that ascorbate enhances the rate of reactivation of the 2-fluoro-glycosyl enzyme, leading to the conclusion that ascorbic acid substitutes for the catalytic base in myrosinase.

Ascorbic Acid↗
PubMed2000

The mechanism of substrate (aglycone) specificity in beta -glucosidases is revealed by crystal structures of mutant maize beta -glucosidase-DIMBOA, -DIMBOAGlc, and -dhurrin complexes.

The mechanism and the site of substrate (i.e., aglycone) recognition and specificity were investigated in maize beta-glucosidase (Glu1) by x-ray crystallography by using crystals of a catalytically inactive mutant (Glu1E191D) in complex with the natural substrate 2-O-beta-d-glucopyranosyl-4-hydroxy-7-methoxy-1,4-benzoxazin-3-one (DIMBOAGlc), the free aglycone DIMBOA, and competitive inhibitor para-hydroxy-S-mandelonitrile beta-glucoside (dhurrin). The structures of these complexes and of the free enzyme were solved at 2.1-, 2.1-, 2.0-, and 2.2-A resolution, respectively. The structural data from the complexes allowed us to visualize an intact substrate, free aglycone, or a competitive inhibitor in the slot-like active site of a beta-glucosidase. These data show that the aglycone moiety of the substrate is sandwiched between W378 on one side and F198, F205, and F466 on the other. Thus, specific conformations of these four hydrophobic amino acids and the shape of the aglycone-binding site they form determine aglycone recognition and substrate specificity in Glu1. In addition to these four residues, A467 interacts with the 7-methoxy group of DIMBOA. All residues but W378 are variable among beta-glucosidases that differ in substrate specificity, supporting the conclusion that these sites are the basis of aglycone recognition and binding (i.e., substrate specificity) in beta-glucosidases. The data also provide a plausible explanation for the competitive binding of dhurrin to maize beta-glucosidases with high affinity without being hydrolyzed.

Amino Acid Sequence↗
PubMed2000

Crystal structure of a cohesin module from Clostridium cellulolyticum: implications for dockerin recognition.

In the assembly of the Clostridium cellulolyticum cellulosome, the multiple cohesin modules of the scaffolding protein CipC serve as receptors for cellulolytic enzymes which bear a dockerin module. The X-ray structure of a type I C. cellulolyticum cohesin module (Cc-cohesin) has been solved using molecular replacement, and refined at 2.0 A resolution. Despite a rather low sequence identity of 32 %, this module has a fold close to those of the two Clostridium thermocellum cohesin (Ct-cohesin) modules whose 3D structures have been determined previously. Cc-cohesin forms a dimer in the crystal, as do the two Ct-cohesins. We show here that the dimer exists in solution and that addition of dockerin-containing proteins dissociates the dimer. This suggests that the dimerization interface and the cohesin/dockerin interface may overlap. The nature of the overall surface and of the dimer interface of Cc-cohesin differ notably from those of the Ct-cohesin modules, being much less polar, and this may explain the species specificity observed in the cohesin/dockerin interaction of C. cellulolyticum and C. thermocellum. We have produced a topology model of a C. cellulolyticum dockerin and of a Cc-cohesin/dockerin complex using homology modeling and available biochemical data. Our model suggests that a special residue pair, already identified in dockerin sequences, is located at the center of the cohesin surface putatively interacting with the dockerin.

Amino Acid Sequence↗