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dna sequence analysis of endoglucanase genes from pseudomonas fluorescens subsp. cellulosa and pseudomonas sp. ncib 8634.the dna of two previously isolated recombinant clones, one from pseudomonas sp. ncib 8634 (= cellvibrio mixtus) (ppc71) and another from pseudomonas fluorescens subsp. cellulosa (ppfc4) that express endoglucanase activity in e. coli was sequenced. plasmid ppc71 had three open reading frames, two of which include portions of plasmid pbr322. the third open reading frame occurs entirely within the pseudomonas dna insert and encodes a protein with a molecular mass of 5845 da. the dna insert in ppfc4 ...19901366996
genes from cellvibrio mixtus encoding beta-1,3 endoglucanase.two genes encoding beta-1,3 glucanase activity were cloned from the gram-negative soil bacterium cellvibrio mixtus. the two clones, designated cwd (cell wall degradation) and lam (laminarin degradation), had distinct endonuclease restriction patterns and encoded enzymes with distinct substrate specificities. the 3.7-kilobase cwd insert encoded an enzyme which degraded yeast cell walls as well as the soluble beta-1,3 glucan laminarin and the insoluble beta-1,3 glucans zymosan and pachyman. the 1. ...19902285322
identification of tandemly repeated type vi cellulose-binding domains in an endoglucanase from the aerobic soil bacterium cellvibrio mixtus.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 ...19989650253
galactomannan hydrolysis and mannose metabolism in cellvibrio mixtus.galactomannan hydrolysis results from the concerted action of microbial endo-mannanases, manosidases and alpha-galactosidases and is a mechanism of intrinsic biological importance. here we report the identification of a gene cluster in the aerobic soil bacterium cellvibrio mixtus encoding enzymes involved in the degradation of this polymeric substrate. the family 27 alpha-galactosidase, termed cmaga27a, preferentially hydrolyse galactose containing polysaccharides. in addition, we have character ...200616842369
expression and evaluation of enzymes required for the hydrolysis of galactomannan.the cost-effective production of bioethanol from lignocellulose requires the complete conversion of plant biomass, which contains up to 30 % mannan. to ensure utilisation of galactomannan during consolidated bioprocessing, heterologous production of mannan-degrading enzymes in fungal hosts was explored. the aspergillus aculeatus endo-β-mannanase (man1) and talaromyces emersonii α-galactosidase (agal) genes were expressed in saccharomyces cerevisiae y294, and the aspergillus niger β-mannosidase ( ...201424888762
characterization of xyn10j, a novel family 10 xylanase from a compost metagenomic library.a gene encoding an extracellular xylanase was cloned from a compost metagenomic library. the xylanase gene, xyn10j, was 1,137 bp in length and was predicted to encode a protein of 378 amino acid residues with a putative signal peptide of 27 amino acid residues. the molecular mass of the mature xyn10j was calculated to be 39,882 da with a pi of 6.09. xyn10j had a motif gvkvhftemdi characteristic of most members of glycosyl hydrolase family 10. the amino acid sequence of xyn10j showed 60.0% identi ...201222215253
novel modular enzymes encoded by a cellulase gene cluster in cellvibrio mixtus.hydrolysis of plant cell wall polysaccharides, a process which is of intrinsic biological and biotechnological importance, requires the concerted action of an extensive repertoire of microbial cellulases and hemicellulases. here, we report the identification of the gene cluster unk16a, rega and cel5b in the aerobic soil bacterium cellvibrio mixtus, encoding a family 16 (cmunk16a) glycoside hydrolase (gh), an arac/xyls transcription activator (cmrega) and a family 5 (cmcel5b) endo-glucanase, resp ...200617005007
probing the structural basis for the difference in thermostability displayed by family 10 xylanases.thermostability is an important property of industrially significant hydrolytic enzymes: understanding the structural basis for this attribute will underpin the future biotechnological exploitation of these biocatalysts. the cellvibrio family 10 (gh10) xylanases display considerable sequence identity but exhibit significant differences in thermostability; thus, these enzymes represent excellent models to examine the structural basis for the variation in stability displayed by these glycoside hyd ...200616762367
characterization of a metagenome-derived halotolerant cellulase.metagenomes of uncultured microorganisms represent a sheer unlimited resource for discovery of novel biocatalysts. here, we report on the biochemical characterisation of a novel, soil metagenome-derived cellulase (endoglucanase), cel5a. the deduced amino acid sequence of cel5a was similar to a family 5, single domain cellulase with no distinct cellulose binding domain from cellvibrio mixtus. the 1092bp orf encoding cel5a was overexpressed in escherichia coli and the corresponding 42.1 kda protei ...200616584799
cloning of a gene cluster from cellvibrio mixtus which codes for cellulase, chitinase, amylase, and pectinase.the soil isolate cellvibrio mixtus uqm2294 degraded a variety of polysaccharides including microcrystalline cellulose. among 6,000 cosmid clones carrying c. mixtus dna, constructed in escherichia coli with phc79, 50 expressed the ability to degrade one or more of the following substrates: carboxymethyl cellulose, chitin, pectin (polygalacturonic acid), cellobiose, and starch. these degradative genes are encoded in a single 94.1-kilobase segment of the c. mixtus genome; a preliminary order of the ...198616347240
reclassification of 'pseudomonas fluorescens subsp. cellulosa' ncimb 10462 (ueda et al. 1952) as cellvibrio japonicus sp. nov. and revival of cellvibrio vulgaris sp. nov., nom. rev. and cellvibrio fulvus sp. nov., nom. rev.'pseudomonas fluorescens subsp. cellulosa' ncimb 10462 has been demonstrated by a polyphasic taxonomic approach to be a member of the genus cellvibrio. 16s rdna sequence analysis suggests that this is the only genus that could accept this specimen. the sequence is 95.5% similar to that of cellvibrio mixtus subsp. mixtus acm 2601t (the type strain of the type species of the genus), which is its closest relation. the genomic dna g + c content was determined to be 53.3 mol%, which is similar to the ...200312710603
molecular cloning, sequencing and expression of the gene encoding a novel chitinase a from a marine bacterium, pseudomonas sp pe2, and its domain structure.the pcha gene encoding chitinase a (pcha) from a pythium porphyrae cell-wall-degrading marine bacterium, pseudomonas sp. pe2, was cloned and characterized. the deduced pcha was a modular enzyme composed of an n-terminal signal peptide, a glycoside hydrolase family 18 catalytic domain that was responsible for the chitinase activity, the chitin-binding domains (chbds), and the carbohydrate-binding modules (cbm). the amino acid sequence of chbd(pcha) was highly conserved in the cbm family 12 that a ...200312655456
isolation and identification of cellulolytic bacteria involved in the degradation of natural cellulosic fibres.in search for bacterial cultures that are able to rapidly degrade cellulosic plant fibres in vitro, 77 cellulolytic strains were isolated from belgian and czech soils after enrichment on flax or sisal fibres as sole sources of carbon. the strains were characterized using fatty acid analysis, and 74 strains were grouped into three major clusters by numerical analysis. the first major cluster contained cellulomonas strains. within this cluster three subclusters could be delineated by principal com ...200010930083
possible roles for a non-modular, thermostable and proteinase-resistant cellulase from the mesophilic aerobic soil bacterium cellvibrio mixtus.the widespread presence of cellulose-binding domains in cellulases from aerobic bacteria and fungi suggests the existence of a strong selective pressure for the retention of these non-catalytic modules. the complete nucleotide sequence of the cellulase gene, cela, from the aerobic soil bacterium cellvibrio mixtus, was determined. it revealed an open reading frame of 1089 bp that encoded a polypeptide, defined as cellulase a (cela), of m(r) 41,548. cela displayed features characteristic of an end ...19979390455
a gene encoding an exo-beta-glucosidase from cellvibrio mixtus.cellvibrio mixtus produces an array of endohydrolytic enzymes involved in the initial phases of beta-glycan polysaccharide degradation in the soil. these enzymes convert complex, high-molecular-weight, insoluble polysaccharides into low-molecular-weight, soluble oligosaccharides which must be further degraded for cellular uptake and catabolism. little is known about the enzymes involved in this latter process in c. mixtus. in this paper we report the cloning of the lam2 gene, which encodes an ex ...19979290063
cloning and characterization of two closely linked cellulase genes from cellvibrio mixtusa 16.5-kb bamhi fragment of the cellvibrio mixtus chromosome was found to direct carboxymethylcellulase, xylanase, and avicel hydrolysis. two closely linked genes were subcloned from this insert. the gene, cmci, was cloned as a 2.7-kb fragment and expressed in escherichia coli. it encoded an enzyme of approximately 74 kda which degraded carboxymethylcellulose and xylan but did not attack the microcrystalline cellulose substrate avicel. a second cellulase capable of degrading avicel, encoded by e ...19968661691
novel cellulose-binding domains, nodb homologues and conserved modular architecture in xylanases from the aerobic soil bacteria pseudomonas fluorescens subsp. cellulosa and cellvibrio mixtus.to test the hypothesis that selective pressure has led to the retention of cellulose-binding domains (cbds) by hemicellulase enzymes from aerobic bacteria, four new xylanase (xyn) genes from two cellulolytic soil bacteria, pseudomonas fluorescens subsp. cellulosa and cellvibrio mixtus, have been isolated and sequenced. pseudomonas genes xyne and xynf encoded modular xylanases (xyle and xylf) with predicted m(r) values of 68,600 and 65000 respectively. xyle contained a glycosyl hydrolase family 1 ...19957492333
fusion of a family 9 cellulose-binding module improves catalytic potential of clostridium thermocellum cellodextrin phosphorylase on insoluble cellulose.clostridium thermocellum cellodextrin phosphorylase (ctcdp), a single-module protein without an apparent carbohydrate-binding module, has reported activities on soluble cellodextrin with a degree of polymerization (dp) from two to five. in this study, ctcdp was first discovered to have weak activities on weakly water-soluble celloheptaose and insoluble regenerated amorphous cellulose (rac). to enhance its activity on solid cellulosic materials, four cellulose binding modules, e.g., cbm3 (type a) ...201121630044
family 6 carbohydrate-binding modules display multiple beta1,3-linked glucan-specific binding interfaces.noncatalytic carbohydrate-binding modules (cbms), which are found in a variety of carbohydrate-degrading enzymes, have been grouped into sequence-based families. cbms, by recruiting their appended enzymes onto the surface of the target substrate, potentiate catalysis particularly against insoluble substrates. family 6 cbms (cbm6s) display unusual properties in that they present two potential ligand-binding sites termed clefts a and b, respectively. cleft b is located on the concave surface of th ...200919751219
insights into the molecular determinants of substrate specificity in glycoside hydrolase family 5 revealed by the crystal structure and kinetics of cellvibrio mixtus mannosidase 5a.the enzymatic hydrolysis of the glycosidic bond is central to numerous biological processes. glycoside hydrolases, which catalyze these reactions, are grouped into families based on primary sequence similarities. one of the largest glycoside hydrolase families is glycoside hydrolase family 5 (gh5), which contains primarily endo-acting enzymes that hydrolyze beta-mannans and beta-glucans. here we report the cloning, characterization, and three-dimensional structure of the cellvibrio mixtus gh5 be ...200415014076
the crystal structure of the family 6 carbohydrate binding module from cellvibrio mixtus endoglucanase 5a in complex with oligosaccharides reveals two distinct binding sites with different ligand specificities.glycoside hydrolases that release fixed carbon from the plant cell wall are of considerable biological and industrial importance. these hydrolases contain non-catalytic carbohydrate binding modules (cbms) that, by bringing the appended catalytic domain into intimate association with its insoluble substrate, greatly potentiate catalysis. family 6 cbms (cbm6) are highly unusual because they contain two distinct clefts (cleft a and cleft b) that potentially can function as binding sites. henshaw et ...200415010454
the family 6 carbohydrate binding module cmcbm6-2 contains two ligand-binding sites with distinct specificities.the microbial degradation of the plant cell wall is an important biological process, representing a major component of the carbon cycle. enzymes that mediate the hydrolysis of this composite structure are modular proteins that contain non-catalytic carbohydrate binding modules (cbms) that enhance catalytic activity. cbms are grouped into sequence-based families, and in a previous study we showed that a family 6 cbm (cbm6) that interacts with xylan contains two potential ligand binding clefts, de ...200415004011
the mechanisms by which family 10 glycoside hydrolases bind decorated substrates.endo-beta-1,4-xylanases (xylanases), which cleave beta-1,4 glycosidic bonds in the xylan backbone, are important components of the repertoire of enzymes that catalyze plant cell wall degradation. the mechanism by which these enzymes are able to hydrolyze a range of decorated xylans remains unclear. here we reveal the three-dimensional structure, determined by x-ray crystallography, and the catalytic properties of the cellvibrio mixtus enzyme xyn10b (cmxyn10b), the most active gh10 xylanase descr ...200414668328
taxonomic study of cellvibrio strains and description of cellvibrio ostraviensis sp. nov., cellvibrio fibrivorans sp. nov. and cellvibrio gandavensis sp. nov.thirty-one cellulolytic bacterial isolates from soils that were phenotypically very similar and phylogenetically highly related to cellvibrio strains were further characterized using a polyphasic taxonomic approach. by using repetitive extragenic palindromic dna-pcr fingerprinting, six different fingerprints could be recognized among the isolates. representative strains and four reference strains of the genus cellvibrio were used for dna-dna hybridization, which yielded eight dna hybridization g ...200312710614
the membrane-bound alpha-glucuronidase from pseudomonas cellulosa hydrolyzes 4-o-methyl-d-glucuronoxylooligosaccharides but not 4-o-methyl-d-glucuronoxylan.the microbial degradation of xylan is a key biological process. hardwood 4-o-methyl-d-glucuronoxylans are extensively decorated with 4-o-methyl-d-glucuronic acid, which is cleaved from the polysaccharides by alpha-glucuronidases. in this report we describe the primary structures of the alpha-glucuronidase from cellvibrio mixtus (c. mixtus glca67a) and the alpha-glucuronidase from pseudomonas cellulosa (p. cellulosa glca67a) and characterize p. cellulosa glca67a. the primary structures of c. mixt ...200212169619
a novel cellvibrio mixtus family 10 xylanase that is both intracellular and expressed under non-inducing conditions.hydrolysis of the plant cell wall polysaccharides cellulose and xylan requires the synergistic interaction of a repertoire of extracellular enzymes. recently, evidence has emerged that anaerobic bacteria can synthesize high levels of periplasmic xylanases which may be involved in the hydrolysis of small xylo-oligosaccharides absorbed by the micro-organism. cellvibrio mixtus, a saprophytic aerobic soil bacterium that is highly active against plant cell wall polysaccharides, was shown to express i ...200010931900
characterization of an endo-1,3(4)-beta-d-glucanase gene from cellvibrio mixtus.an endo-1,3(4)-beta-d-glucanase gene (cwd2) of cellvibrio mixtus encoding laminarinase activity was cloned on a 3.9-kb psti fragment. the cwd2 enzyme, extracted from recombinant escherichia coli, degraded both beta-1,3 glucans and beta-1,3-1,4 mixed-linkage glucans, was endohydrolytic and so conformed to the enzyme class 3.2.1.6. the ph and temperature optima of the enzyme were approximately 7 and 40 degrees c respectively. the m(r) of specifically labelled cwd2 was approximately 34,000. this ge ...19938339916
hydrolysis of wheat flour arabinoxylan, acid-debranched wheat flour arabinoxylan and arabino-xylo-oligosaccharides by β-xylanase, α-l-arabinofuranosidase and β-xylosidase.a range of α-l-arabinofuranosyl-(1-4)-β-d-xylo-oligosaccharides (axos) were produced by hydrolysis of wheat flour arabinoxylan (wax) and acid debranched arabinoxylan (adwax), in the presence and absence of an axh-d3 α-l-arabinofuranosidase, by several gh10 and gh11 β-xylanases. the structures of the oligosaccharides were characterised by gc-ms and nmr and by hydrolysis by a range of α-l-arabinofuranosidases and β-xylosidase. the axos were purified and used to characterise the action patterns of ...201525723624
characterization of a xylanase-producing cellvibrio mixtus strain j3-8 and its genome analysis.cellvibrio mixtus strain j3-8 is a gram-negative, xylanase-producing aerobic soil bacterium isolated from giant snails in singapore. it is able to produce up to 10.1 u ml(-1) of xylanase, which is comparable to xylanase production from known bacterial and fungal strains. genome sequence analysis of strain j3-8 reveals that the assembled draft genome contains 5,171,890 bp with a g + c content of 46.66%, while open reading frame (orf) annotations indicate a high density of genes encoding glycoside ...201525994900
draft genome sequence of a xylanase-producing bacterial strain, cellvibrio mixtus j3-8.the xylanase-producing bacterial strain cellvibrio mixtus j3-8 was isolated from grassland giant snails. the draft genome of strain j3-8 comprises 5,171,890 bp in 152 contigs with a g+c content of 46.66%. this is the first genome report about this bacterial species.201425502674
factor g utilizes a carbohydrate-binding cleft that is conserved between horseshoe crab and bacteria for the recognition of beta-1,3-d-glucans.in the horseshoe crab, the recognition of beta-1,3-d-glucans by factor g triggers hemolymph coagulation. factor g contains a domain of two tandem xylanase z-like modules (z1-z2), each of which recognizes beta-1,3-d-glucans. to gain an insight into the recognition of beta-1,3-d-glucans from a structural view point, recombinants of z1-z2, the c-terminal module z2, z2 with a cys to ala substitution (z2a), and its tandem repeat z2a-z2a were characterized. z2 and z1-z2, but not z2a and z2a-z2a, forme ...200919710471
cloning and functional characterization of endo-β-1,4-glucanase gene from metagenomic library of vermicompost.in the vermicomposting of paper mill sludge, the activity of earthworms is very dependent on dietetic polysaccharides including cellulose as energy sources. most of these polymers are degraded by the host microbiota and considered potentially important source for cellulolytic enzymes. in the present study, a metagenomic library was constructed from vermicompost (vc) prepared with paper mill sludge and dairy sludge (fresh sludge, fs) and functionally screened for cellulolytic activities. eighteen ...201323812813
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