| reclassification of thermomonospora and microtetraspora. | almost complete 16s rrna sequences from seven thermomonospora strains, thermomonospora curvata, thermomonospora formosensis, thermomonospora fusca, thermomonospora mesophila, thermomonospora chromogena, thermomonospora alba and thermomonospora mesouviformis (a synonym of thermomonospora alba) were determined and subjected to phylogenetic analysis together with the sequences from all the representative members of the suborder streptosporangineae. on the basis of phylogenetic, chemotaxonomic and p ... | 1998 | 9731279 |
| effects of noncatalytic residue mutations on substrate specificity and ligand binding of thermobifida fusca endocellulase cel6a. | the availability of a high-resolution structure of the thermobifida fusca endocellulase cel6a catalytic domain makes this enzyme ideal for structure-based efforts to engineer cellulases with high activity on native cellulose. in order to determine the role of conserved, noncatalytic residues in cellulose hydrolysis, 14 mutations of six conserved residues in or near the cel6a active-site cleft were studied for their effects on catalytic activity, substrate specificity, processivity and ligand-bin ... | 2000 | 10601873 |
| a celr mutation affecting transcription of cellulase genes in thermobifida fusca. | biosynthesis of extracellular cellulases in the cellulose-degrading actinomycete thermobifida fusca is controlled by a transcriptional regulator, celr, and cellobiose, which acts as an inducer interfering with the celr-dna interaction. we report the identification and characterization of a mutation in the celr gene that changes ala(55) in the hinge helix of celr to thr. the wild-type and mutant celr genes were cloned in escherichia coli, and their protein products were characterized. the celr mu ... | 2000 | 10613893 |
| site-directed mutation of noncatalytic residues of thermobifida fusca exocellulase cel6b. | fifteen mutant genes in six loop residues and eight mutant genes in five conserved noncatalytic active site residues of thermobifida fusca cel6b were constructed, cloned and expressed in escherichia coli or streptomyces lividans. the mutant enzymes were assayed for catalytic activity on carboxymethyl cellulose (cmc), swollen cellulose (sc), filter paper (fp), and bacterial microcrystalline cellulose (bmcc) as well as cellotetraose, cellopentaose, and 2, 4-dinitrophenyl-beta-d-cellobioside. they ... | 2000 | 10824094 |
| cloning, expression and characterization of a family 48 exocellulase, cel48a, from thermobifida fusca. | the gene for a 104-kda exocellulase, cel48a, formerly e6, was cloned from thermobifida fusca into escherichia coli and streptomyces lividans. the dna sequence revealed a type ii cellulose-binding domain at the n-terminus, followed by a fniii-like domain and ending with a glycosyl hydrolase family 48 catalytic domain. the enzyme and catalytic domain alone were each expressed in and purified from s. lividans and had very low catalytic activity on swollen cellulose, carboxymethyl cellulose, bacteri ... | 2000 | 10931180 |
| cloning and biochemical characterization of bglc, a beta-glucosidase from the cellulolytic actinomycete thermobifida fusca. | an operon, bglabc, that encodes two sugar permeases and a beta-glucosidase was cloned from a cellulolytic actinomycete, thermobifida fusca, into escherichia coli and sequenced. the bglc gene encoding an intracellular beta-glucosidase (beta-d-glucoside glucohydrolase, ec 3.2.1.21) belonging to glycosyl hydrolase family 1 was subcloned and expressed in e. coli. the purified enzyme (mw 53,407 da; pi 4.69) hydrolyzed substrates containing both beta 1 --> 4 and beta 1 --> 2 glycosidic bonds, and was ... | 2001 | 11178732 |
| surface structures of new and lesser known species of thermobifida as revealed by scanning electron microscopy. | surface structures of representatives of the genus thermobifida were examined by scanning electron microscopy. spores formed at the tips of multibranched sporophores initially resembled short sausages; then, upon maturation, they gradually built up their typical ovoid shape. characteristic differences were observed between t. cellulolytica strain tb108 and t. fusca strains tm51. the spores of tb108 were larger (0.8x 1.3 microm) than those of tm51 (0.6 x 1.1 microm) in consequence of the more thi ... | 2001 | 11426855 |
| phylogenetic diversity of thermophilic actinomycetes and thermoactinomyces spp. isolated from mushroom composts in korea based on 16s rrna gene sequence analysis. | forty one strains isolated from 21 samples of various mushroom composts in korea were analyzed by 16s rrna gene sequencing to investigate the phylogenetic diversity of thermophilic actinomycetes. the 25 strains of thermophilic actinomycete isolates were related to the five genera, pseudonocardia, saccharomonospora, saccharopolyspora, streptomyces, and thermobifida, within the order actinomycetales, and 16 strains were classified into the genus thermoactinomyces within the family bacillaceae. mos ... | 2001 | 11506914 |
| 4-methyl-7-thioumbelliferyl-beta-d-cellobioside: a fluorescent, nonhydrolyzable substrate analogue for cellulases. | the kinetics of cellulose binding and hydrolysis by cellulases is not well understood except at steady-state conditions. for use in studies of cellulase pre-steady-state and steady-state kinetics, we have prepared 4-methyl-7-thioumbelliferyl-beta-d-cellobioside (mus-cb), a ground-state nonhydrolyzable analogue of the fluorescent cellulase substrate 4-methylumbelliferyl-beta-d-cellobioside (mu-cb). mus-cb is not hydrolyzed by the catalytic domain of cellulase e1 from acidothermus cellulolyticus u ... | 2002 | 11914092 |
| synergism in binary mixtures of thermobifida fusca cellulases cel6b, cel9a, and cel5a on bmcc and avicel. | in an earlier binding study conducted in our laboratory using thermobifida fusca cellulases cel6b, cel9a, and cel5a (formally thermomonosporafusca e3, e4, and e5), it was observed that binding capacities for these three cellulases were 18-30 times higher on bmcc than on avicel. these results stimulated an interest in how the difference in accessibility between the two cellulosic substrates would affect synergism observed with cellulase mixtures. to explore the impact of substrate accessibility o ... | 2002 | 12049205 |
| thermobifida cellulolytica sp. nov., a novel lignocellulose-decomposing actinomycete. | four actinomycete strains, isolated from the overheated region of manure compost, were assigned to the genus thermobifida on the basis of morphological, physiological and biochemical characteristics. all strains produced single, ovoid, heat-sensitive spores on dichotomically branched aerial hyphae. on the basis of chemotaxonomic traits, these isolates showed strong affinity towards members of the genus thermobifida. cell-wall analysis revealed the presence of meso-diaminopimelic acid, but no oth ... | 2002 | 12148627 |
| cooperative and competitive binding in synergistic mixtures of thermobifida fusca cellulases cel5a, cel6b, and cel9a. | synergism between cellulases facilitates efficient hydrolysis of microcrystalline cellulose. we hypothesize that the effects of synergism, observed as enhanced extents of hydrolysis, are related to cellulase binding to the substrate in mixtures. in this study, direct measurements of bound concentrations of fluorescence-labeled t. fusca cel5a, cel6b, and cel9a on bacterial microcrystalline cellulose were used to study binding behaviors of cellulases in binary component reactions. the accuracy of ... | 2002 | 12153310 |
| binding mechanisms for thermobifida fusca cel5a, cel6b, and cel48a cellulose-binding modules on bacterial microcrystalline cellulose. | the family ii cellulose-binding modules (cbm) from thermobifida fusca cel5a and cel48a were cloned in the escherichia coli/streptomyces shuttle vector pd730, and the plasmids were transformed into streptomyces lividans tkm31. cbm(cel5a), and cbm(cel48a), cbm(cel6b) were expressed and purified from s. lividans. the molecular masses were determined by mass spectrometry, and the values were 10595 +/- 2, 10915 +/- 2, and 11291 +/- 2 da for cbm(cel5a), cbm(cel6b), and cbm(cel48a), respectively. three ... | 2002 | 12325146 |
| transcript analysis of genes encoding a family 61 endoglucanase and a putative membrane-anchored family 9 glycosyl hydrolase from phanerochaete chrysosporium. | phanerochaete chrysosporium cellulase genes were cloned and characterized. the cel61a product was structurally similar to fungal endoglucanases of glycoside hydrolase family 61, whereas the cel9a product revealed similarities to thermobifida fusca cel9a (e4), an enzyme with both endo- and exocellulase characteristics. the fungal cel9a is apparently a membrane-bound protein, which is very unusual for microbial cellulases. transcript levels of both genes were substantially higher in cellulose-grow ... | 2002 | 12406778 |
| serpins in prokaryotes. | members of the serpin (serine proteinase inhibitor) superfamily have been identified in higher multicellular eukaryotes (plants and animals) and viruses but not in bacteria, archaea, or fungi. thus, the ancestral serpin and the origin of the serpin inhibitory mechanism remain obscure. in this study we characterize 12 serpin-like sequences in the genomes of prokaryotic organisms, extending this protein family to all major branches of life. notably, these organisms live in dramatically different e ... | 2002 | 12411597 |
| cloning and heterologous expression of a beta-d-mannosidase (ec 3.2.1.25)-encoding gene from thermobifida fusca tm51. | thermobifida fusca tm51, a thermophilic actinomycete isolated from composted horse manure, was found to produce a number of lignocellulose-degrading hydrolases, including endoglucanases, exoglucanases, endoxylanases, beta-xylosidases, endomannanases, and beta-mannosidases, when grown on cellulose or hemicellulose as carbon sources. beta-mannosidases (ec 3.2.1.25), although contributing to the hydrolysis of hemicellulose fractions, such as galacto-mannans, constitute a lesser-known group of the l ... | 2003 | 12676668 |
| computational and experimental studies of the catalytic mechanism of thermobifida fusca cellulase cel6a (e2). | mutagenesis experiments suggest that asp79 in cellulase cel6a (e2) from thermobifida fusca has a catalytic role, in spite of the fact that this residue is more than 13 a from the scissile bond in models of the enzyme-substrate complex built upon the crystal structure of the protein. this suggests that there is a substantial conformational shift in the protein upon substrate binding. molecular mechanics simulations were used to investigate possible alternate conformations of the protein bound to ... | 2003 | 12676981 |
| the non-catalytic amino acid asp446 is essential for enzyme activity of the modular endocellulase cel9 from myxobacter sp. al-1. | the modular endocellulase cel9 of the bicistronic operon cel9-cel48 of myxobacter sp. al-1 shares not only amino acid sequence similarity but also biochemical properties similar to those of thermobifida fusca endo/exocellulase e4. amino acid alignments of a t. fusca e4 cellulase subfamily of family 9 cellulases revealed that asp(446) of myxobacter sp. al-1 cel9, a putatively noncatalytic residue, is highly conserved in one of the catalytic domains of this subfamily. directed mutagenesis of resid ... | 2003 | 12732983 |
| corn fiber hydrolysis by thermobifida fusca extracellular enzymes. | thermobifida fusca was grown on cellulose (solka-floc), xylan or corn fiber and the supernatant extracellular enzymes were concentrated. sds gels showed markedly different protein patterns for the three different carbon sources. activity assays on a variety of synthetic and natural substrates showed major differences in the concentrated extracellular enzyme activities. these crude enzyme preparations were used to hydrolyze corn fiber, a low-value biomass byproduct of the wet milling of corn. app ... | 2003 | 12743765 |
| [teichoic acids in the cell walls of microbispora mesophila ac-1953t and thermobifida fusca ac-1952t]. | the cell walls of microbispora mesophila strain ac-1953t (the family streptosporangiaceae) and thermobifida fusca ac-1952t (the family nocardiopsiceae) were found to contain teichoic acids of a poly(glycerol phosphate) nature. the teichoic acid of m. mesophila (formerly thermomonospora mesophila) represents a poly(glycerol phosphate) containing 5% of substituent 2-acetamido-2-deoxy-alpha-galactosaminyl residues. the teichoic acid of such kind was found in actinomycetes for the first time. the ce ... | 2003 | 12751242 |
| cloning, expression and characterization of a family-74 xyloglucanase from thermobifida fusca. | thermobifida fusca xyloglucan-specific endo-beta-1,4-glucanase (xeg)74 and the xeg74 catalytic domain (cd) were cloned, expressed in escherichia coli, purified and characterized. this enzyme has a glycohydrolase family-74 cd that is a specific xyloglucanase followed by a family-2 carbohydrate binding module at the c terminus. the michaelis constant (km) and maximal rate (vmax) values for hydrolysis of tamarind seed xyloglucan (tamxg) are 2.4 micro m and 966 micro mol xyloglucan oligosaccharides ... | 2003 | 12846842 |
| numerical phenetic classification of clinically significant aerobic sporoactinomycetes and related organisms. | clinically significant aerobic sporoactinomycetes, notably agents of mycetoma, were examined for a balanced set of unit characters and the resultant data analysed using standard numerical taxonomic procedures. all save two of the one hundred and seventy three tested strains were assigned to three multimembered cluster-groups, which encompassed sixteen major (4-7 strains), ten minor (2-3 strains) and forty single membered clusters, in an analysis based on the simple matching coefficient and unwei ... | 2003 | 12906361 |
| quantitative determination of noncovalent binding interactions using automated nanoelectrospray mass spectrometry. | electrospray ionization mass spectrometry (esi-ms) has proven to be an extremely powerful tool for studying biomolecular structures and noncovalent interactions. here we report a method using a fully automated, chip-based nanoesi-ms system to determine the dissociation constants (kd) for the complexes of two different proteins with their ligands. the automated nanoelectrospray system, consisting of the nanomate and esi chip, serves functionally as a combination of autosampler and nanoelectrospra ... | 2003 | 12964745 |
| binding and reversibility of thermobifida fusca cel5a, cel6b, and cel48a and their respective catalytic domains to bacterial microcrystalline cellulose. | the binding and reversibility of thermobifida fusca intact cel5a, cel5b, and cel48a and their corresponding catalytic domains (cds) to bacterial microcrystalline cellulose (bmcc) were studied at 5 degrees c. the binding of the intact cellulases and of corresponding cds to bmcc was irreversible in all regions: langmuir binding (region i), interstice penetration (region ii), and interstice saturation (region iii). the three cellulose binding domains (cbms) bind reversibly in "region i" although th ... | 2003 | 12966571 |
| horizontal gene transfer from eukarya to bacteria and domain shuffling: the alpha-amylase model. | alpha-amylases are present in all kingdoms of the living world. despite strong conservation of the tertiary structure, only a few amino acids are conserved in interkingdom comparisons. animal alpha-amylases are characterized by several typical motifs and biochemical properties. a few cases of such alpha-amylases have been previously reported in some eubacterial species. we screened the bacterial genomes available in the sequence databases for new occurrences of animal-like alpha-amylases. three ... | 2004 | 14704857 |
| integration of computer modeling and initial studies of site-directed mutagenesis to improve cellulase activity on cel9a from thermobifida fusca. | cellulases are a complex group of enzymes that are fundamental for the degradation of amorphous and crystalline cellulose in lignocellulosic material. unfortunately, cellulases have a low catalytic efficiency on their substrates when compared to similar enzymes such as amylases, which has led to a strong interest in improving their activities. thermobifida fusca secretes six cellulose degrading enzymes: two exo- and three endocellulases and an endo/exocellulase cel9a (formerly called e4). cel9a ... | 2004 | 15054213 |
| studies of thermobifida fusca plant cell wall degrading enzymes. | i have been studying the thermobifida fusca cellulose degrading proteins for the past 25 years. in this period, we have purified and characterized the six extracellular cellulases and an intracellular beta- glucosidase used by t. fusca for cellulose degradation, cloned and sequenced the structural genes encoding these enzymes, and helped to determine the 3-dimensional structures of two of the cellulase catalytic domains. this research determined the mechanism of a novel class of cellulase, famil ... | 2004 | 15073875 |
| synergistic activity of paenibacillus sp. bp-23 cellobiohydrolase cel48c in association with the contiguous endoglucanase cel9b and with endo- or exo-acting glucanases from thermobifida fusca. | cellobiohydrolase cel48c from paenibacillus sp. bp-23, an enzyme displaying limited activity on most cellulosic substrates, was assayed for activity in the presence of other bacterial endo- or exocellulases. significant enhanced activity was observed when cel48c was incubated in the presence of paenibacillus sp. bp-23 endoglucanase cel9b or thermobifida fusca cellulases cel6a and cel6b, indicating that cel48c acts synergistically with them. maximum synergism rates on bacterial microcrystalline c ... | 2004 | 15236244 |
| cloning, expression and identification of a new trehalose synthase gene from thermobifida fusca genome. | a new open reading frame in thermobifida fusca sequenced genome was identified to encode a new trehalose synthase, annotated as "glycosidase" in the genbank database, by bioinformatics searching and experimental validation. the gene had a length of 1830 bp with about 65% gc content and encoded for a new trehalose synthase with 610 amino acids and deduced molecular weight of 66 kd. the high gc content seemed not to affect its good expression in e. coli bl21 in which the target protein could accou ... | 2004 | 15248022 |
| kinetic studies of thermobifida fusca cel9a active site mutant enzymes. | thermobifida fusca cel9a-90, an unusual family 9 enzyme, is a processive endoglucanase containing a catalytic domain closely linked to a family 3c cellulose binding domain (cel9a-68) followed by a fibronectin iii-like domain and a family 2 cellulose binding domain. to study its catalytic mechanism, 12 mutant genes with changes in five conserved residues of cel9a-68 were constructed, cloned, and expressed in escherichia coli. the purified mutant enzymes were assayed for their activities on (carbo ... | 2004 | 15274620 |
| recombinant expression and enzymatic characterization of pttcel9a, a kor homologue from populus tremula x tremuloides. | pttcel9a is a membrane-bound, family 9 glycosyl hydrolase from populus tremula x tremuloides that is upregulated during secondary cell wall synthesis. the catalytic domain of pttcel9a, delta(1-105)pttcel9a, was purified, and its activity was compared to tfcel9a and tfcel9b from thermobifida fusca. since aromatic amino acids involved in substrate binding at subsites -4, -3, and -2 are missing in pttcel9a, the activity of tfcel9a mutant enzymes w256s, w209a, and w313g was also investigated. delta( ... | 2004 | 15287736 |
| crystal structure of a baeyer-villiger monooxygenase. | flavin-containing baeyer-villiger monooxygenases employ nadph and molecular oxygen to catalyze the insertion of an oxygen atom into a carbon-carbon bond of a carbonylic substrate. these enzymes can potentially be exploited in a variety of biocatalytic applications given the wide use of baeyer-villiger reactions in synthetic organic chemistry. the catalytic activity of these enzymes involves the formation of two crucial intermediates: a flavin peroxide generated by the reaction of the reduced fla ... | 2004 | 15328411 |
| cloning, characterization and phylogenetic relationships of cel5b, a new endoglucanase encoding gene from thermobifida fusca. | thermobifida fusca, a thermophilic, aerobic, cellulolytic bacterium has a highly complex cellulase system comprising three endoglucanases, two exoglucanases and one processive endoglucanase. zymogram analysis indicated that additional cellulases may exist in t. fusca strain tm51, therefore a tm51 expression library was prepared in streptomyces lividans tk24 and screened for hydrolases. a new endoglucanase gene, named tf cel5b, was identified. heterologous cel5b, produced in s. lividans, had temp ... | 2004 | 15378527 |
| optimized pathway selection in intraresidual triple-resonance experiments. | an optimized intraresidual pulse sequence element with better sensitivity and suppression of sequential cross peaks is presented. concatenation of three magnetization transfer delays allows their independent setting, in accordance with the relaxation properties of the individual spins, without concomitantly prolonging the pulse sequence. additionally, implementations of the scheme to hnca, hncacb, and the trosy based triple-resonance experiments are proposed. the feasibility of the new element w ... | 2004 | 15388087 |
| the high resolution crystal structure of a native thermostable serpin reveals the complex mechanism underpinning the stressed to relaxed transition. | serpins fold into a native metastable state and utilize a complex conformational change to inhibit target proteases. an undesirable result of this conformational flexibility is that most inhibitory serpins are heat sensitive, forming inactive polymers at elevated temperatures. however, the prokaryote serpin, thermopin, from thermobifida fusca is able to function in a heated environment. we have determined the 1.8 a x-ray crystal structure of thermopin in the native, inhibitory conformation. a st ... | 2004 | 15590653 |
| discovery of a thermostable baeyer-villiger monooxygenase by genome mining. | baeyer-villiger monooxygenases represent useful biocatalytic tools, as they can catalyze reactions which are difficult to achieve using chemical means. however, only a limited number of these atypical monooxygenases are available in recombinant form. using a recently described protein sequence motif, a putative baeyer-villiger monooxygenase (bvmo) was identified in the genome of the thermophilic actinomycete thermobifida fusca. heterologous expression of the respective protein in escherichia col ... | 2004 | 15599520 |
| characterization of a new extracellular hydrolase from thermobifida fusca degrading aliphatic-aromatic copolyesters. | the paper describes the purification, biochemical characterization, sequence determination, and classification of a novel thermophilic hydrolase from thermobifida fusca (tfh) which is highly active in hydrolyzing aliphatic-aromatic copolyesters. the secretion of the extracellular enzyme is induced by the presence of aliphatic-aromatic copolyesters but also by adding several other esters to the medium. the hydrophobic enzyme could be purified applying a combination of (nh(4))so(4)-precipitation, ... | 2005 | 15638529 |
| purification and characterization of thermobifida fusca xylanase 10b. | thermobifida fusca grows well on cellulose and xylan, and produces a number of cellulases and xylanases. the gene encoding a previously unstudied endoxylanase, xyl10b, was overexpressed in e. coli, and the protein was purified and characterized. mature xyl10b is a 43-kda glycohydrolase with a short basic domain at the c-terminus. it has moderate thermostability, maintaining 50% of its activity after incubation for 16 h at 62 degrees c, and is most active between ph 5 and 8. xyl10b is produced by ... | 2004 | 15644898 |
| a novel thermostable hemoglobin from the actinobacterium thermobifida fusca. | the gene coding for a hemoglobin-like protein (tf-trhb) has been identified in the thermophilic actinobacterium thermobifida fusca and cloned in escherichia coli for overexpression. the crystal structure of the ferric, acetate-bound derivative, was obtained at 2.48 a resolution. the three-dimensional structure of tf-trhb is similar to structures reported for the truncated hemoglobins from mycobacterium tuberculosis and bacillus subtilis in its central domain. the complete lack of diffraction pat ... | 2005 | 16098200 |
| a single amino acid substitution enhances the catalytic activity of family 11 xylanase at alkaline ph. | random mutagenesis of the gene encoding family 11 xylanase was used to obtain alkalophilic mutants. the catalytic domain of the chimeric enzyme stx15, which was constructed from streptomyces lividans xylanase b and thermobifida fusca xylanase a, was mutated using error-prone pcr and screened for halo formation on dye-linked xylan plates and activity toward soluble xylan. a positive mutant, m1011, was isolated, and it was found that mutation a49v was responsible for the alkalophilicity of the mut ... | 2005 | 16116276 |
| crystal structure of thermobifida fusca endoglucanase cel6a in complex with substrate and inhibitor: the role of tyrosine y73 in substrate ring distortion. | endoglucanase cel6a from thermobifida fusca hydrolyzes the beta-1,4 linkages in cellulose at accessible points along the polymer. the structure of the catalytic domain of cel6a from t. fusca in complex with a nonhydrolysable substrate analogue that acts as an inhibitor, methylcellobiosyl-4-thio-beta-cellobioside (glc(2)-s-glc(2)), has been determined to 1.5 a resolution. the glycosyl unit in subsite -1 was sterically hindered by tyr73 and forced into a distorted (2)s(o) conformation. in the enzy ... | 2005 | 16185060 |
| beta-d-glucosidase reaction kinetics from isothermal titration microcalorimetry. | the cellobiase activities of nine thermal stable mutants of thermobifida fusca bglc were assayed by isothermal titration microcalorimetry (itc). the mutations were previously generated using random mutagenesis and identified by high-temperature screening as imparting improved thermal stability to the beta-d-glucosidase enzyme. analysis of the substrate-saturation curves obtained by itc for the wild-type enzyme and the nine thermally stabilized mutants revealed that the wild type and all the muta ... | 2005 | 16269126 |
| phenolic extraction from apple peel by cellulases from thermobifida fusca. | with the optimization of the pretreatment conditions for the crude thermobifida fusca cellulase activity and phenolic release from apple peel, we focused on the activity of individual purified cellulase related to the antioxidant activity. the overall phenolic release was significantly increased in a synergistic manner with combined pretreatment, not with individual pretreatment such as boiling, acid, and pectinase treatment. approximately 60 mg of reducing sugar equivalent were produced per g o ... | 2005 | 16302777 |
| new model substrates for enzymes hydrolysing polyethyleneterephthalate and polyamide fibres. | recently the potential of enzymes for surface hydrophilisation and/or functionalisation of polyethyleneterephthalate (pet) and polyamide (pa) has been discovered. however, there was no correlation between enzyme class/activity (e.g. esterase, lipase, cutinase) and surface hydrolysis of these polymers and consequently no simple assay to estimate this capability. enzymes active on the model substrates bis (benzoyloxyethyl) terephthalate and adipic acid bishexyl-amide, were also capable of increasi ... | 2006 | 16624419 |
| signature proteins that are distinctive characteristics of actinobacteria and their subgroups. | the actinobacteria constitute one of the main phyla of bacteria. presently, no morphological and very few molecular characteristics are known which can distinguish species of this highly diverse group. in this work, we have analyzed the genomes of four actinobacteria (viz. mycobacterium leprae tn, leifsonia xyli subsp. xyli str. ctcb07, bifidobacterium longum ncc2705 and thermobifida fusca yx) to search for proteins that are unique to actinobacteria. our analyses have identified 233 actinobacter ... | 2006 | 16670965 |
| production of a recombinant polyester-cleaving hydrolase from thermobifida fusca in escherichia coli. | the hydrolase (thermobifida fusca hydrolase; tfh) from t. fusca was produced in escherichia coli as fusion protein using the ompa leader sequence and a his(6) tag. productivity could be raised more than 100-fold. both batch and fed-batch cultivations yield comparable cell specific productivities whereas volumetric productivities differ largely. in the fed-batch cultivations final rtfh concentrations of 0.5 g l(-1) could be achieved. in batch cultivations the generated rtfh is translocated to the ... | 2006 | 16770590 |
| increase of the hydrophilicity of polyethylene terephthalate fibres by hydrolases from thermomonospora fusca and fusarium solani f. sp. pisi. | treatment of polyethylene terephthalate fibres with hydrolase preparations from thermomonospora (thermobifida) fusca and fusarium solani f. sp. pisi resulted in an increase of the hydrophilicity of the fibres determined by measurement of their dyeing behaviour with reactive dyes and their water absorption ability. reflectance spectrometry of treated fibres dyed with a reactive dye showed that the colour became more intense corresponding to an increase of hydroxyl groups on the fibre surfaces and ... | 2006 | 16791721 |
| codon optimized thermobifida fusca hydrolase secreted by bacillus megaterium. | production and secretion of a 28,172 da hydrolase from thermobifida fusca (tfh) in bacillus megaterium ms941 and wh323 was investigated in shake flask and ph controlled bioreactors. successful production of heterologous tfh was achieved by adapting the original tfh gene to the optimal codon usage of b. megaterium. a codon adaption index close to one was reached. the codon optimized tfh was cloned into an open reading frame with dna sequence for the n-terminal signal peptide of b. megaterium lipa ... | 2007 | 16948171 |
| new genus-specific primers for the pcr identification of novel isolates of the genus streptomonospora. | the halophilic actinomycete genus streptomonospora, forming a distinct branch in the 16s rrna gene phylogenetic tree adjacent to the genera nocardiopsis and thermobifida, was first proposed by cui et al. to accommodate the species type streptomonospora salina. during a biodiversity and taxonomic study on halophilic filamentous actinomycetes from a saline lake in western china, numerous new halophilic actinomycetes strains were isolated. to confirm whether they are members of the genus streptomon ... | 2006 | 16958850 |
| characterization of a thermobifida fusca beta-1,3-glucanase (lam81a) with a potential role in plant biomass degradation. | thermobifida fusca is a filamentous soil bacterium that plays a major role in the breakdown of plant biomass. in this paper, we report the cloning, expression, purification, and characterization of the t. fusca enzyme, lam81a. the carbohydrate active enzymes database (http://afmb.cnrs-mrs.fr/cazy/) indicates that lam81a belongs to a relatively uncharacterized family of beta-1,3-glucanases, family gh-81 [coutinho, p. m., and henrissat, b. (1999) in recent advances in carbohydrate bioengineering ( ... | 2006 | 17115704 |
| genome sequence and analysis of the soil cellulolytic actinomycete thermobifida fusca yx. | thermobifida fusca is a moderately thermophilic soil bacterium that belongs to actinobacteria. it is a major degrader of plant cell walls and has been used as a model organism for the study of secreted, thermostable cellulases. the complete genome sequence showed that t. fusca has a single circular chromosome of 3,642,249 bp predicted to encode 3,117 proteins and 65 rna species with a coding density of 85%. genome analysis revealed the existence of 29 putative glycoside hydrolases in addition to ... | 2007 | 17209016 |
| cloning and characterization of a maltotriose-producing alpha-amylase gene from thermobifida fusca. | the gene (tfa), encoding a maltotriose-producing alpha-amylase from thermobifida fusca ntu22, was cloned, sequenced and expressed in escherichia coli. the gene consists of 1,815 base pairs and encodes a protein of 605 amino acids. the base composition of the tfa coding sequence is 69% g+c and the protein has a predicted pi value of 5.5. the deduced amino acid sequence of the tfa amylase exhibited a high degree of similarity with amylases from thermomonospora curvata and streptomyces amylases. th ... | 2007 | 17211634 |
| enhanced cutinase production with thermobifida fusca by two-stage ph control strategy. | a mutant of thermobifida fusca atcc 27730 was used for cutinase production. acetate was the most suitable carbon source for cell growth and cutinase production compared with others. the ph was one of the most important factors affecting cutinase yield and productivity. batch cutinase fermentations by mutant thermobifida fusca wsh04 at various ph values ranging from 7.0 to 7.9 were studied. based on the effects of different ph values on the specific cell growth rate and specific cutinase formatio ... | 2007 | 17309045 |
| a tomato endo-beta-1,4-glucanase, slcel9c1, represents a distinct subclass with a new family of carbohydrate binding modules (cbm49). | a critical structural feature of many microbial endo-beta-1,4-glucanases (egases, or cellulases) is a carbohydrate binding module (cbm), which is required for effective crystalline cellulose degradation. however, cbms are absent from plant egases that have been biochemically characterized to date, and accordingly, plant egases are not generally thought to have the capacity to degrade crystalline cellulose. we report the biochemical characterization of a tomato egase, solanum lycopersicum cel8 (s ... | 2007 | 17322304 |
| processivity, substrate binding, and mechanism of cellulose hydrolysis by thermobifida fusca cel9a. | thermobifida fusca cel9a-90 is a processive endoglucanase consisting of a family 9 catalytic domain (cd), a family 3c cellulose binding module (cbm3c), a fibronectin iii-like domain, and a family 2 cbm. this enzyme has the highest activity of any individual t. fusca enzyme on crystalline substrates, particularly bacterial cellulose (bc). mutations were introduced into the cd or the cbm3c of cel9a-68 using site-directed mutagenesis. the mutant enzymes were expressed in escherichia coli; purified; ... | 2007 | 17369336 |
| production of xylooligosaccharides from xylans by extracellular xylanases from thermobifida fusca. | xylooligosaccharides are produced for use as a valuable food sweetener or additive. they have many beneficial biomedical and health effects. in this study, a process for producing xylooligosaccharides from lignocellulolytic agricultural waste was developed. bagasse, corncob, wheat bran, and peanut shell were used as carbon sources for production of xylanolytic enzymes from thermobifida fusca ntu22. when using bagasse as the carbon source, the xylanolytic enzymes that simultaneously accumulated i ... | 2007 | 17432873 |
| mesophile versus thermophile: insights into the structural mechanisms of kinetic stability. | obtaining detailed knowledge of folding intermediate and transition state (ts) structures is critical for understanding protein folding mechanisms. comparisons between proteins adapted to survive extreme temperatures with their mesophilic homologs are likely to provide valuable information on the interactions relevant to the unfolding transition. for kinetically stable proteins such as alpha-lytic protease (alphalp) and its family members, their large free energy barrier to unfolding is central ... | 2007 | 17543987 |
| a novel chimera: the "truncated hemoglobin-antibiotic monooxygenase" from streptomyces avermitilis. | novel chimeric proteins made of a globin domain fused with a "cofactor free" monooxygenase domain have been identified within the streptomyces avermitilis and frankia sp. genomes by means of bioinformatics methods. structure based sequence alignments show that the globin domains of both proteins can be unambiguously assigned to the truncated hemoglobin family, in view of the striking similarity to the truncated hemoglobins from mycobacterium tuberculosis, thermobifida fusca and bacillus subtilis ... | 2007 | 17574781 |
| proteomic and transcriptomic analysis of extracellular proteins and mrna levels in thermobifida fusca grown on cellobiose and glucose. | thermobifida fusca secretes proteins that carry out plant cell wall degradation. using two-dimensional electrophoresis, the extracellular proteome of t. fusca grown on cellobiose was compared to that of cells grown on glucose. extracellular proteins, the expression of which is induced by cellobiose, mainly are cellulases and cellulose-binding proteins. other major extracellular proteins induced by cellobiose include a xylanase (xyl10a) and two unknown proteins, the c-terminal regions of which ar ... | 2007 | 17601791 |
| effects of differing temperature management on development of actinobacteria populations during composting. | actinobacteria are believed to play a major role in organic matter degradation and humification processes in composts. in this study, the effects of different temperature regimes on the succession of actinobacteria populations during composting were investigated in a laboratory reactor. phospholipid fatty acid (plfa) was used to investigate quantitative changes in the overall microbial biomass and community structure, and in the size of actinobacteria populations. qualitative changes were determ ... | 2007 | 17683913 |
| automated docking to explore subsite binding by glycoside hydrolase family 6 cellobiohydrolases and endoglucanases. | cellooligosaccharides were computationally docked using autodock into the active sites of the glycoside hydrolase family 6 enzymes hypocrea jecorina (formerly trichoderma reesei) cellobiohydrolase and thermobifida fusca endoglucanase. subsite -2 exerts the greatest intermolecular energy in binding beta-glucosyl residues, with energies progressively decreasing to either side. cumulative forces imparting processivity exerted by these two enzymes are significantly less than by the equivalent glycos ... | 2007 | 17724729 |
| expression of thermostable microbial cellulases in the chloroplasts of nicotine-free tobacco. | an inexpensive source of active cellulases is critical to efficient and cost-effective conversion of lignocellulosic biomass to ethanol. transgenic plants expressing foreign cellulases are potential sources of cellulases for biomass conversion. a number of foreign proteins have been reported to accumulate to high levels when the transgene is incorporated into the chloroplast genome rather than into the nuclear genome. we developed plastid transformation vectors carrying two thermobifida fusca th ... | 2007 | 17765995 |
| multiple display of catalytic modules on a protein scaffold: nano-fabrication of enzyme particles. | self assembly is a prerequisite for fabricating nanoscale structures. here we present a new fusion protein based on the stress-responsive homo-oligomeric protein, sp1. this ring-shaped protein is a highly stable homododecamer, which can be potentially utilized to self-assemble different modules and enzymes in a predicted and oriented manner. for that purpose, a cohesin module (a component of the bacterial cellulosome) was selected, its gene fused in-frame to sp1, and the fusion protein was expre ... | 2007 | 17826857 |
| effect of different carbon sources on central metabolic fluxes and the recombinant production of a hydrolase from thermobifida fusca in bacillus megaterium. | the recombinant bacillus megaterium strain wh323 was employed for the inducible production and secretion of recombinant thermobifida fusca hydrolase (tfh). continuous cultivations were carried out in a chemostat using either glucose or pyruvate as sole carbon source. a remarkable increase of produced tfh was detected for the pyruvate-dependent cultivation compared to glucose-dependent growth. estimation of intracellular carbon fluxes through the central metabolism for both growth conditions usin ... | 2007 | 17826861 |
| enhancement of the activity and alkaline ph stability of thermobifida fusca xylanase a by directed evolution. | directed evolution has been used to enhance the catalytic activity and alkaline ph stability of thermobifida fusca xylanase a, which is one of the most thermostable xylanases. under triple screened traits of activity, alkaline ph stability and thermostability, through two rounds of random mutagenesis using dna shuffling, a mutant 2tfxa98 with approximately 12-fold increased k(cat)/k(m) and 4.5-fold decreased k(m) compared with its parent was obtained. moreover, the alkaline ph stability of 2tfxa ... | 2008 | 18292971 |
| chitin binding by thermobifida fusca cellulase catalytic domains. | cellulose is a linear homopolymer of beta 1-4 linked glucose residues. chitin is similar to cellulose in structure, and can be described as cellulose with the hydroxyl group on the c2 carbon replaced by an acetylamine group. both cellulose and chitin form tightly packed, extensively hydrogen-bonded micro-fibrils. up to now, binding of cellulase catalytic domains (cds) to chitin has not been reported. in this article, binding of the cds of thermobifida fusca cel6a, cel6b, cel48a, cel5a, and cel9a ... | 2008 | 18306418 |
| kinetic mechanism of phenylacetone monooxygenase from thermobifida fusca. | phenylacetone monooxygenase (pamo) from thermobifida fusca is a fad-containing baeyer-villiger monooxygenase (bvmo). to elucidate the mechanism of conversion of phenylacetone by pamo, we have performed a detailed steady-state and pre-steady-state kinetic analysis. in the catalytic cycle ( k cat = 3.1 s (-1)), rapid binding of nadph ( k d = 0.7 microm) is followed by a transfer of the 4( r)-hydride from nadph to the fad cofactor ( k red = 12 s (-1)). the reduced pamo is rapidly oxygenated by mole ... | 2008 | 18321069 |
| enzymatic hydrolysis of ptt polymers and oligomers. | oligomers and polymers (film, fabrics) of the linear aromatic polyester poly(trimethylene terephthalate) (ptt) were treated with polyesterases from thermomyces lanuginosus, penicillium citrinum, thermobifida fusca and fusarium solani pisi. the cutinase from t. fusca was found to release the highest amounts of hydrolysis products from ptt materials and was able to open and hydrolyse a cyclic ptt dimer according to rp-hplc-uv detection. in contrast, the lipase from t. lanuginosus also showed activ ... | 2008 | 18405994 |
| regulation and characterization of thermobifida fusca carbohydrate-binding module proteins e7 and e8. | e7, a single domain family 33 cellulose binding module (cbm) protein, and e8, a non-catalytic, three-domain protein consisting of a family 33 cbm, a fniii domain, followed by a family 2 cbm, were cloned, expressed, purified, and characterized. western blots showed that e7 and e8 were induced and secreted when thermobifida fusca was grown on cellobiose, solka floc, switchgrass, or alfalfa as well as on beta-1,3 linked glucose molecules such as laminaribiose or pachyman. e8 bound well to alpha- an ... | 2008 | 18553392 |
| immobilization of cellulose fibrils on solid substrates for cellulase-binding studies through quantitative fluorescence microscopy. | cellulases, enzymes capable of depolymerizing cellulose polymers into fermentable sugars, are essential components in the production of bioethanol from lignocellulosic materials. given the importance of these enzymes to the evolving biofuel industry considerable research effort is focused on understanding the interaction between cellulases and cellulose fibrils. this manuscript presents a method that addresses challenges that must be overcome in order to study such interactions through high-reso ... | 2008 | 18563846 |
| conversion of thermobifida fusca free exoglucanases into cellulosomal components: comparative impact on cellulose-degrading activity. | cellulosomes are multi-enzyme complexes produced by certain anaerobic bacteria that exhibit efficient degradation of plant cell wall polysaccharides. to understand their enhanced levels of hydrolysis, we are investigating the effects of converting a free-cellulase system into a cellulosomal one. to achieve this end, we are replacing the cellulose-binding module of the native cellulases, produced by the aerobic bacterium thermobifida fusca, with a cellulosome-derived dockerin module of establishe ... | 2008 | 18582975 |
| dgge and t-rflp analysis of bacterial succession during mushroom compost production and sequence-aided t-rflp profile of mature compost. | the amount of button mushroom (agaricus bisporus) harvested from compost is largely affected by the microbial processes taking place during composting and the microbes inhabiting the mature compost. in this study, the microbial changes during the stages of this specific composting process were monitored, and the dominant bacteria of the mature compost were identified to reveal the microbiological background of the favorable properties of the heat-treated phase ii mushroom compost. 16s ribosomal ... | 2009 | 18654815 |
| identification and characterization of bacterial cutinase. | cutinase, which exists in both fungi and bacteria, catalyzes the cleavage of the ester bonds of cutin. fungal cutinases have been extensively studied, however, reports on bacterial cutinases have been limited due to the lack of knowledge concerning the identity of their open reading frames. in the present study, the cutinase from thermobifida fusca was induced by cutin and purified to homogeneity by following p-nitrophenyl butyrate hydrolyzing activity. peptide mass fingerprinting analysis of th ... | 2008 | 18658138 |
| thermobifida halotolerans sp. nov., isolated from a salt mine sample, and emended description of the genus thermobifida. | the actinomycete strain yim 90462t was isolated from a salt mine sample collected from yunnan province, south-west china. comparative 16s rrna gene sequence analysis demonstrated that the isolate, which had sequence similarities of more than 97 % to thermobifida alba, thermobifida fusca and thermobifida cellulosilytica, was a candidate member of the genus thermobifida. subsequently, a polyphasic approach was used to clarify the taxonomic position of strain yim 90462t. its diagnostic diamino acid ... | 2008 | 18676463 |
| [cutinase production from short-chain organic acids by thermobifida fusca]. | we studied cutinase production from short-chain organic acids by thermobifida fusca wsh03-11 to evaluate the possibility of converting municipal sludge to high value-added products. the optimum organic acid (8.0 g/l) and nitrogen source (1.5 g/l) concentrations were determined by the single factor experiments with butyric acid, propionic acid and acetic acid as the carbon sources. when lactic acid was used as the carbon source, the optimum organic acid (3.0 g/l) and nitrogen source (1.0 g/l) con ... | 2008 | 18724703 |
| comparison of organic matter degradation and microbial community during thermophilic composting of two different types of anaerobic sludge. | changes in organic matter degradation and microbial communities during thermophilic composting were compared using two different types of anaerobic sludge, one from mesophilic methane fermentation, containing a high concentration of proteins (s-sludge), and the other from thermophilic methane fermentation, containing high concentrations of lipids and fibers (k-sludge). the difference in the organic matter degradation rate corresponded to the difference in the organic matter constituents; the co( ... | 2009 | 18762416 |
| structure elucidation and biosynthesis of fuscachelins, peptide siderophores from the moderate thermophile thermobifida fusca. | bacteria belonging to the order actinomycetales have proven to be an important source of biologically active and often therapeutically useful natural products. the characterization of orphan biosynthetic gene clusters is an emerging and valuable approach to the discovery of novel small molecules. analysis of the recently sequenced genome of the thermophilic actinomycete thermobifida fusca revealed an orphan nonribosomal peptide biosynthetic gene cluster coding for an unknown siderophore natural ... | 2008 | 18832174 |
| interaction between the cbm of cel9a from thermobifida fusca and cellulose fibers. | molecular docking and molecular dynamics (md) simulations were used to investigate the binding of a cellodextrin chain in a crystal-like conformation to the carbohydrate-binding module (cbm) of cel9a from thermobifida fusca. the fiber was found to bind to the cbm in a single and well-defined configuration in-line with the catalytic cleft, supporting the hypothesis that this cbm plays a role in the catalysis by feeding the catalytic domain (cd) with a polysaccharide chain. the results also expand ... | 2009 | 18853469 |
| macroamphiphilic components of thermophilic actinomycetes: identification of lipoteichoic acid in thermobifida fusca. | the cell envelopes of gram-positive bacteria contain structurally diverse membrane-anchored macroamphiphiles (lipoteichoic acids and lipoglycans) whose functions are poorly understood. since regulation of membrane composition is an important feature of adaptation to life at higher temperatures, we have examined the nature of the macroamphiphiles present in the thermophilic actinomycetes thermobifida fusca and rubrobacter xylanophilus. following hot-phenol-water extraction and purification by hyd ... | 2009 | 18931132 |
| high-level bacterial cellulase accumulation in chloroplast-transformed tobacco mediated by downstream box fusions. | the thermobifida fusca cel6a gene encoding an endoglucanase was fused to three different downstream box (db) regions to generate cel6a genes with 14 amino acid fusions. the db-cel6a fusions were inserted into the tobacco (nicotiana tabacum cv. samsun) chloroplast genome for protein expression. accumulation of cel6a protein in transformed tobacco leaves varied over approximately two orders of magnitude, dependent on the identity of the db region fused to the cel6a open reading frame (orf). additi ... | 2009 | 18973281 |
| heterologous expression of polyhydroxyalkanoate depolymerase from thermobifida sp. in pichia pastoris and catalytic analysis by surface plasmon resonance. | a polyhydroxyalkanote depolymerase gene from thermobifida sp. isolate bcc23166 was cloned and expressed as a c-terminal his(6)-tagged fusion in pichia pastoris. primary structure analysis revealed that the enzyme phaz-th is a member of a proposed new subgroup of scl-pha depolymerase containing a proline-serine repeat linker. phaz-th was expressed as two glycosylated forms with apparent molecular weights of 61 and 70 kda, respectively. the enzyme showed esterase activity toward p-nitrophenyl alka ... | 2009 | 19002683 |
| expression patterns of cel5a-cel5b, two endoglucanase encoding genes of thermobifida fusca. | expression patterns of cel5a and cel5b, two endoglucanase encoding genes of thermobifida fusca were compared by quantitative real-time pcr. with avicel as carbon source the transcript level of cel5a continuously increased until the 10th hour of incubation and then a sharp decrease was observed, whereas cel5b presented a slow constitutive expression on this substrate. when the microcrystalline cellulose powder mn300 was used as the inducing carbon source, the expression patterns of the two genes ... | 2008 | 19130751 |
| site-directed mutagenesis to probe catalysis by a thermobifida fusca beta-1,3-glucanase (lam81a). | thermobifida fuscalam81a is a single domain family-81 beta-1,3-endoglucanase, but no structure is known for this family. site-directed mutagenesis of 14 conserved residues chosen from sequence alignments was used to identify those with critical roles in catalysis, binding or substrate specificity. mutant enzymes were assayed for their ability to bind and hydrolyze substrates with various glycosyl linkages. residues d422, e499 and e503 were candidates for the catalytic acid or catalytic base, and ... | 2009 | 19435780 |
| cloning and functional analysis of cis-prenyltransferase from thermobifida fusca. | cis-prenyltransferase catalyzes the synthesis of z,e-mixed prenyl diphosphates by a condensation of isopentenyl diphosphate to an allylic diphosphate. a novel gene encoding a cis-prenyltransferase is cloned from thermobifida fusca. it showed a unique substrate specificity accepting dimethylallyl diphosphate as a shortest allylic substrate, and synthesizes polyprenyl products up to c(70). | 2009 | 19447338 |
| the absence of an identifiable single catalytic base residue in thermobifida fusca exocellulase cel6b. | thermobifida fusca exocellulase cel6b acts by an inverting hydrolysis mechanism; however, the catalytic acid and base residues for this enzyme have not been confirmed. site-directed mutagenesis and kinetic studies were used to show that asp274 is the catalytic acid, which is consistent with what is found for other members of family-6 glycoside hydrolases; however, a single catalytic base was not identified. mutation of all putative catalytic base residues, within 6 a of the -1/+1 glucose subsite ... | 2009 | 19523117 |
| structural and functional characterizations of ssgb, a conserved activator of developmental cell division in morphologically complex actinomycetes. | ssga-like proteins (salps) are a family of homologous cell division-related proteins that occur exclusively in morphologically complex actinomycetes. we show that ssgb, a subfamily of salps, is the archetypal salp that is functionally conserved in all sporulating actinomycetes. sporulation-specific cell division of streptomyces coelicolor ssgb mutants is restored by introduction of distant ssgb orthologues from other actinomycetes. interestingly, the number of septa (and spores) of the complemen ... | 2009 | 19567872 |
| the microbial signature of aerosols produced during the thermophilic phase of composting. | the microbial diversity of bioaerosols released during operational activities at composting plants is poorly understood. identification of bacteria and fungi present in such aerosols is the prerequisite for the definition of microbial indicators that could be used in dispersal and exposure studies. | 2010 | 19602015 |
| enzymatic surface hydrolysis of poly(ethylene terephthalate) and bis(benzoyloxyethyl) terephthalate by lipase and cutinase in the presence of surface active molecules. | a lipase from thermomyces lanuginosus and cutinases from thermobifida fusca and fusarium solani hydrolysed poly(ethylene terephthalate) (pet) fabrics and films and bis(benzoyloxyethyl) terephthalate (3pet) endo-wise as shown by maldi-tof-ms, lc-uvd/ms, cationic dyeing and xps analysis. due to interfacial activation of the lipase in the presence of triton x-100, a seven-fold increase of hydrolysis products released from 3pet was measured. in the presence of the plasticizer n,n-diethyl-2-phenylace ... | 2009 | 19616594 |
| the anabaena sensory rhodopsin transducer defines a novel superfamily of prokaryotic small-molecule binding domains. | the anabaena sensory rhodopsin transducer (asrt) is a small protein that has been claimed to function as a signaling molecule downstream of the cyanobacterial sensory rhodopsin. however, orthologs of asrt have been detected in several bacteria that lack rhodopsin, raising questions about the generality of this function. using sequence profile searches we show that asrt defines a novel superfamily of beta-sandwich fold domains. through contextual inference based on domain architectures and predic ... | 2009 | 19682383 |
| single molecule analysis of bacterial polymerase chain reaction products in submicrometer fluidic channels. | laser induced fluorescence in submicrometer fluidic channels was used to characterize the synthesis of polymerase chain reaction (pcr) products from a model bacterial system in order to explore the advantages and limitations of on chip real time single molecule pcr analysis. single oligonucleotide universal bacterial primers and pcr amplicons from the 16s rdna of thermobifida fusca (325 bp) were directly detected at all phases of the reaction with low sample consumption and without post-amplific ... | 2007 | 19693361 |
| influence of culture aeration on the cellulase activity of thermobifida fusca. | currently, one of the hurdles hindering efficient production of cellulosic biofuel is the recalcitrant nature of cellulose to hydrolysis. a wide variety of cellulase enzymes are found natively in microorganisms that can potentially be used to effectively hydrolyze cellulose to fermentable sugars. in this study, phenomenological and mechanistic parameters affecting cellulase activity were studied using the moderately thermophilic, aerobic, and cellulolytic microorganism thermobifida fusca. two ma ... | 2010 | 19697023 |
| processivity, synergism, and substrate specificity of thermobifida fusca cel6b. | a relationship between processivity and synergism has not been reported for cellulases, although both characteristics are very important for hydrolysis of insoluble substrates. mutation of two residues located in the active site tunnel of thermobifida fusca exocellulase cel6b increased processivity on filter paper. surprisingly, mixtures of the cel6b mutant enzymes and t. fusca endocellulase cel5a did not show increased synergism or processivity, and the mutant enzyme which had the highest proce ... | 2009 | 19734341 |
| novel alkali-stable, cellulase-free xylanase from deep-sea kocuria sp. mn22. | a novel xylanase gene, kxyn, was cloned from kocuria sp. mn22, a bacteria isolated from the deep sea of the east pacific. kxyn consists of 1,170 bp and encodes a protein of 390 amino acids that shows the highest identity (63%) with a xylanase from thermobifida fusca yx. the mature protein with a molecular mass of approximately 40 kda was expressed in escherichia coli bl21 (de3). the recombinant kxyn displayed its maximum activity at 55 degrees and at ph 8.5. the km, vmax, and kcat values of kxyn ... | 2009 | 19809242 |
| effect of linker length and dockerin position on conversion of a thermobifida fusca endoglucanase to the cellulosomal mode. | we have been developing the cellulases of thermobifida fusca as a model to explore the conversion from a free cellulase system to the cellulosomal mode. three of the six t. fusca cellulases (endoglucanase cel6a and exoglucanases cel6b and cel48a) have been converted in previous work by replacing their cellulose-binding modules (cbms) with a dockerin, and the resultant recombinant "cellulosomized" enzymes were incorporated into chimeric scaffolding proteins that contained cohesin(s) together with ... | 2009 | 19820154 |
| gene cloning, expression, and characterization of a thermostable xylanase from nesterenkonia xinjiangensis cctcc aa001025. | an endo-beta-1,4-xylanase-encoding gene, xyn11nx, was cloned from nesterenkonia xinjiangensis cctcc aa001025 and expressed in escherichia coli. the gene encoded a 192-amino acid polypeptide and a putative 50-amino acid signal peptide. the deduced amino acid sequence exhibited a high degree of similarity with the xylanases from streptomyces thermocyaneoviolaceus (68%) and thermobifida fusca (66%) belonging to glycoside hydrolase family 11. after purification to homogeneity, the recombinant xyn11n ... | 2010 | 19838860 |
| a novel expression system for intracellular production and purification of recombinant affinity-tagged proteins in aspergillus niger. | a set of different integrative expression vectors for the intracellular production of recombinant proteins with or without affinity tag in aspergillus niger was developed. target genes can be expressed under the control of the highly efficient, constitutive pkia promoter or the novel sucrose-inducible promoter of the beta-fructofuranosidase (suca) gene of a. niger in the presence or absence of alternative carbon sources. all expression plasmids contain an identical multiple cloning sequence that ... | 2010 | 19908039 |
| a robust and extracellular heme-containing peroxidase from thermobifida fusca as prototype of a bacterial peroxidase superfamily. | dyp-type peroxidases comprise a novel superfamily of heme-containing peroxidases which is unrelated to the superfamilies of known peroxidases and of which only a few members have been characterized in some detail. here, we report the identification and characterization of a dyp-type peroxidase (tfudyp) from the thermophilic actinomycete thermobifida fusca. biochemical characterization of the recombinant enzyme showed that it is a monomeric, heme-containing, thermostable, and tat-dependently expo ... | 2010 | 19967355 |
| unusual proximal heme pocket geometry in the deoxygenated thermobifida fusca: a combined spectroscopic investigation. | the spectroscopic properties of the deoxygenated truncated hemoglobin from the actinobacterium thermobifida fusca have been investigated by means of extended x-ray absorption fine structure (exafs), x-ray absorption near edge structure (xanes), and near infrared spectroscopies both at room and cryogenic temperatures. at room temperature the near infrared charge transfer band iii occurs at 772nm, a value that is unusually high for a canonical deoxygenated hemoglobin species, and can only be found ... | 2010 | 20022418 |
| expression of thermobifida fusca thermostable raw starch digesting alpha-amylase in pichia pastoris and its application in raw sago starch hydrolysis. | a gene encoding the thermostable raw starch digesting alpha-amylase in thermobifida fusca ntu22 was amplified by pcr, sequenced and cloned into pichia pastoris x-33 host strain using the vector pgapzalphaa, allowing constitutive expression and secretion of the protein. recombinant expression resulted in high levels of extracellular amylase production, as high as 510 u/l in the hinton flask culture broth. the purified amylase showed a single band at about 65 kda by sds-polyacrylamide gel electrop ... | 2010 | 20039188 |
| development and application of a pcr-targeted gene disruption method for studying celr function in thermobifida fusca. | thermobifida fusca is a high-g+c-content, thermophilic, gram-positive soil actinobacterium with high cellulolytic activity. in t. fusca, celr is thought to act as the primary regulator of cellulase gene expression by binding to a 14-bp inverted repeat [5'-(t)gggagcgctccc(a)] that is upstream of many known cellulase genes. previously, the ability to study the roles and regulation of cellulase genes in t. fusca has been limited largely by a lack of established genetic engineering methods for t. fu ... | 2010 | 20097808 |