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Don L. Crawford - One of the best experts on this subject based on the ideXlab platform.
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cloning sequencing and characterization of two clustered cellulase encoding genes cels1 and cels2 from Streptomyces Viridosporus t7a and their expression in escherichia coli
Actinomycetologica, 2000Co-Authors: Suganthi Ramachandran, Timothy S. Magnuson, Don L. CrawfordAbstract:A recombinant clone, pBLP, containing a 4.1kb fragment of Streptomyces Viridosporus T7A chromosomal DNA was shown to confer endoglucanase activity in Escherichia coli cells. Further subcloning and sequence analysis revealed two co-transcribed Open Reading Frames (ORF) with high sequence similarities to other Streptomyces endoglucanase-encoding genes. A signal peptide, a catalytic domain and cellulose binding domain were identified within ORF1 (celS1). The amino acid sequence of ORF2 (celS2) showed similarity with cellulose binding protein (p40) of Streptomyces halstedii. The hydrophobic analysis of CelSl revealed that it belongs to the family H cellu-lase catalytic domain. The rare TTA codon encoding leucine was found in the signal sequence of both the cellu-lase ORFs, indicating translational dependence on the bldA gene product, a cognate tRNA responsible for translating the TTA codons. The presence of 14 bp inverted repeats in the 5’-end of these genes, was consistent with the highly conserved positive regulatory structure found in other endoglucanase genes.
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cloning sequencing and characterization of the xylanase encoding gene svxa from Streptomyces Viridosporus t7a and its expression in escherichia coli
Actinomycetologica, 2000Co-Authors: Deepa N Kanhiyur, Timothy S. Magnuson, Don L. CrawfordAbstract:A xylanase-encoding gene from lignocellulolytic Streptomyces Viridosporus T7A was isolated, expressed, cloned and sequenced. The gene was isolated by screening an Escherichia coli cosmid genomic library of S. Viridosporus T7A genomic using a substrate overlay method. One xylanase expressing cosmid clone contained a 30 kb DNA insert. The xylanase gene was subsequently subcloned on a 2 kb PstI fragment and sequenced. Analysis of the nucleotide sequence revealed a 915 bp open reading frame coding for a 329 amino acid protein with a calculated molecular weight of 33,550 Da. This gene, designated svxA (Genbank accession number AF198618), shows high sequence homology to genes belonging to xylanase family G.
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Use of azo dye ligand chromatography for the partial purification of a novel extracellular peroxidase from Streptomyces Viridosporus T7A.
Applied microbiology and biotechnology, 1998Co-Authors: N. S. Burke, Don L. CrawfordAbstract:Crude peroxidase preparations from the lignocellulose-degrading actinomycete, Streptomyces Viridosporus T7A, were shown to decolorize several azo dye isomers and showed a correlation of dye structure to degradability similar to that shown by fungal Mn-peroxidase, an enzyme not previously described in actinomycetes. Addition of the heme-peroxidase inhibitor KCN did not significantly change the ability of the T7A enzyme(s) to decompose the dyes. These results suggest that T7A may produce a Mn- or other peroxidase with similar substrate specificity to Mn-peroxidase. Affinity chromatography using immobilized azo dye isomers was used for purifying peroxidases from T7A. A significantly purified peroxidase preparation was obtained irrespective of the azo dye used. In comparison, concanavalin A lectin affinity chromatography showed very poor binding and resolution for T7A peroxidases. Azo dye affinity purification gave preparations sufficiently purified to allow amino acid microsequencing for two of the bound proteins. N-terminal amino acid sequences were found to share significant homology with a fungal Mn-peroxidase and actinomycete cellulases.
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Cloning of clustered Streptomyces Viridosporus T7A lignocellulose catabolism genes encoding peroxidase and endoglucanase and their extracellular expression in Pichia pastoris.
Canadian journal of microbiology, 1998Co-Authors: Latha Thomas, Don L. CrawfordAbstract:A 4.1-kb fragment of chromosomal DNA from the lignocellulose-decomposing actinomycete Streptomyces Viridosporus T7A was previously found to encode a lignin peroxidase gene. However, when cloned into Escherichia coli in pBSKS+, peroxidase activity was not expressed. When cloned in pIJ702 in Streptomyces lividans, the gene was expressed in a peroxidase positive background, owing to the production by S. lividans of its own extracellular peroxidases. To circumvent these problems, the DNA was cloned into the commercial expression vector pIC9 for extracellular expression in the yeast Pichia pastoris. Yeast transformants, however, expressed two activities, extracellular peroxidase and an extracellular endoglucanase. The enzymes were not expressed by the yeast cells alone or by yeast cells with pIC9 without the insert. Expression of the enzymes by only those transformants expressing the 4.1-kb DNA was confirmed by Western blot analyses, by nondenaturing activity gel staining, and by spectrophotometric enzyme assa...
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Purification and characterization of an alkaline xylanase from Streptomyces Viridosporus T7A
Enzyme and Microbial Technology, 1997Co-Authors: Timothy S. Magnuson, Don L. CrawfordAbstract:Streptomyces Viridosporus T7A produces a large amount of extracellular xylanase activity. We have isolated and purified an alkaline xylanase from culture supernatants of this organism. Purification was achieved by treatment of the culture supernatant with Q-Sepharose, concentration of the unbound proteins by ultrafiltration, fractionation of these proteins by Sephadex G-75 gel filtration chromatography, followed by Rotofor preparative isoelectric focusing. The xylanase has a molecular mass of 59 kDa as determined by capillary electrophoresis. The purified xylanase has a pI of 10.2–10.5, a pH optimum of 7.0–8.0, and a temperature optimum of 65–70°C. The xylanase is strongly inhibited by Hg2+, Cu2+, and Fe3+ metal ions. The purified protein shows activity on birchwood and oat spelt xylans, but does not exhibit activity toward galactomannan, arabinogalactan, lichenan, or carboxymethylcellulose. This enzyme could have potential uses in biotechnology applications due to its high pH and temperature optima and unique substrate specificity.
D. L. Crawford - One of the best experts on this subject based on the ideXlab platform.
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Bioprocessing of lignite coals using reductive microorganisms. Final technical report, September 30, 1988--March 29, 1992
1992Co-Authors: D. L. CrawfordAbstract:In order to convert lignite coals into liquid fuels, gases or chemical feedstock, the macromolecular structure of the coal must be broken down into low molecular weight fractions prior to further modification. Our research focused on this aspect of coal bioprocessing. We isolated, characterized and studied the lignite coal-depolymerizing organisms Streptomyces Viridosporus T7A, Pseudomonas sp. DLC-62, unidentified bacterial strain DLC-BB2 and Gram-positive Bacillus megaterium strain DLC-21. In this research we showed that these bacteria are able to solubilize and depolymerize lignite coals using a combination of biological mechanisms including the excretion of coal solublizing basic chemical metabolites and extracellular coal depolymerizing enzymes.
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Bioprocessing of lignite coals using reductive microorganisms
1992Co-Authors: D. L. CrawfordAbstract:In order to convert lignite coals into liquid fuels, gases or chemical feedstock, the macromolecular structure of the coal must be broken down into low molecular weight fractions prior to further modification. Our research focused on this aspect of coal bioprocessing. We isolated, characterized and studied the lignite coal-depolymerizing organisms Streptomyces Viridosporus T7A, Pseudomonas sp. DLC-62, unidentified bacterial strain DLC-BB2 and Gram-positive Bacillus megaterium strain DLC-21. In this research we showed that these bacteria are able to solubilize and depolymerize lignite coals using a combination of biological mechanisms including the excretion of coal solublizing basic chemical metabolites and extracellular coal depolymerizing enzymes.
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Comparison of extracellular peroxidase- and esterase-deficient mutants of Streptomyces Viridosporus T7A.
Applied and environmental microbiology, 1992Co-Authors: T S Magnuson, D. L. CrawfordAbstract:Peroxidase-deficient mutants of the lignin-degrading bacterium Streptomyces Viridosporus T7A were screened for their production of acid-precipitable polymeric lignin, extracellular peroxidases and esterases, and immunoreactivities against a polyclonal antibody produced against electrophoretically purified peroxidase isoform P3 of wild-type S. Viridosporus. The mutants showed diminished abilities to solubilize lignin and produce acid-precipitable polymeric lignin. Their peroxidase activities were decreased, and their esterase production patterns were altered. Western immunoblots demonstrated that the mutants produced proteins immunologically reactive with the antibody, but with different mobilities from those of wild-type proteins. These findings confirm a direct role for peroxidases in lignin solubilization. They also indicate a possible role for esterases.
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Synthesis and properties of lignin peroxidase from Streptomyces Viridosporus T7A.
Applied biochemistry and biotechnology, 1991Co-Authors: S J Loudha, Roger A. Korus, D. L. CrawfordAbstract:The production of lignin peroxidase by Streptomyces Viridosporus T7A was studied in shake flasks and under aerobic conditions in a 7.5-L batch fermentor. Lignin peroxidase synthesis was found to be strongly affected by catabolite repression. Lignin peroxidase was a non-growth-associated, secondary metabolite. The maximum lignin peroxidase activity was 0.064 U/mL at 36 h. In order to maximize lignin peroxidase activity, optimal conditions were determined. The optimal incubation temperature, pH, and substrate (2,4-dichlorophenol) concentration for the enzyme assays were 45 degrees C, 6, and 3 mM, respectively. Stability of lignin peroxidase was determined at 37, 45, and 60 degrees C, and over the pH range 4-9.
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Effects of bacterial lignin peroxidase on organic carbon mineralization in soil, using recombinant Streptomyces strains
Canadian Journal of Microbiology, 1991Co-Authors: Zemin Wang, B. Bleakley, D. L. Crawford, Timothy S. Magnuson, Greg HertelAbstract:To study the effects of bacterial lignin peroxidase ALip-P3 of Streptomyces Viridosporus T7A on the rate of organic carbon turnover in soil, purified lignin peroxidase, with and without addition of...
Roland Blondeau - One of the best experts on this subject based on the ideXlab platform.
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Effect of light on germinating spores of Streptomyces Viridosporus
FEMS microbiology letters, 1999Co-Authors: Marlène Imbert, Roland BlondeauAbstract:Our study showed that the effect of light on germinating spores of Streptomyces was variable: some species were indifferent, whereas others, such as Streptomyces Viridosporus, displayed a marked inhibition of CFU numbers on growth medium. A special study with S. Viridosporus showed that light only had an impact during the first few hours of spore incubation. The effects of scavengers of toxic forms of oxygen and of photosensitizers, along with the oxidative stress of illuminated spores evidenced by the superoxide dismutase levels, suggested that light and oxygen had a combined action.
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Isolation of soil Streptomyces strains capable of degrading humic acids and analysis of their peroxidase activity.
FEMS Microbiology Ecology, 1995Co-Authors: Khadija Dari, Max Béchet, Roland BlondeauAbstract:Fifteen Streptomyces strains capable of decolorizing humic acids in presence of glucose were isolated from soil samples using the dilute suspension technique and spread on agar plates. Six strains, displaying a significant and stable activity, were selected for further characterization. Some features of these isolates (carbon source utilization, enzyme production, antibiotic resistance) were compared with those of the reference strain Streptomyces Viridosporus ATCC 39115. Degradation properties studied in batch cultures at pH 7.0 showed that the catabolic activity on humic acids was generally stimulated by incubation with 100% oxygen and was cell surface-associated. Peroxidase activity from cell-free extracts was analysed by using the oxidation of N,N,N′,N′-tetramethyl-phenylene-diamine. PAGE analysis revealed the existence of two major types of peroxidases (molecular mass: about 39.2 and 61.6 kDa), dividing the strains into two groups. The role of cell surface-associated peroxidase activity in the breakdown of humic acids is discussed.
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Catabolic activity on humic acids of Streptomyces Viridosporus grown under oxygen
Canadian Journal of Microbiology, 1993Co-Authors: C. Yanze Kontchou, Max Béchet, Roland BlondeauAbstract:When Streptomyces Viridosporus was grown in batch culture on mineral salts – glucose medium and under oxygen, aerial mycelium formation and sporulation were suppressed. Under these conditions, biodegradation of soil or synthetic humic acids (expressed as the rate of decolorization of growth medium and the loss of radioactivity of labeled melanoidin, respectively) increased. Concurrently, both the catabolic activity of cells (expressed as peroxidase production, measured by the oxidation of N,N,N′,N′-tetramethyl-1,4-phenylenediamine as substrate) and the excretion of siderophores were stimulated.Key words: humic acid biodegradation, melanoidin mineralization, Streptomyces Viridosporus, peroxidase, siderophore.
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13C NMR spectroscopic analysis of soil humic acids recovered after incubation with some white rot fungi and actinomycetes
Soil Biology and Biochemistry, 1992Co-Authors: Bertrand Dehorter, Clotaire Yanze Kontchou, Roland BlondeauAbstract:Abstract Culture media containing soil humic acids were inoculated with either white-rot fungi (Phanerochaete chrysosporium and Trametes versicolor) or actinomycetes (Streptomyces Viridosporus and Streptomyces sp.) and incubated until a loss of about 30% of humic acids had occurred in the growth medium. The humic acids were recovered by the use of a metal chelate affinity procedure and studied by 13C NMR spectroscopy in order to collect more information about the mechanism of attack on humic acids. The spectra resulting from fungal activity were the most modified; however, no important changes in the main constituents of these compounds and no increase in carboxyl content were recorded. The results suggest an unusual degradation process.
Jennifer R. Davis - One of the best experts on this subject based on the ideXlab platform.
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Genome Sequence of Streptomyces Viridosporus Strain T7A ATCC 39115, a Lignin-Degrading Actinomycete.
Genome announcements, 2013Co-Authors: Jennifer R. Davis, Lynne Goodwin, Hazuki Teshima, Chris Detter, Roxanne Tapia, Cliff Han, Marcel Huntemann, Chia-lin Wei, James Han, Amy ChenAbstract:We announce the availability of the genome sequence of Streptomyces Viridosporus strain T7A ATCC 39115, a plant biomass- degrading actinomycete. This bacterium is of special interest because of its capacity to degrade lignin, an underutilized compo- nent of plants in the context of bioenergy. It has a full complement of genes for plant biomass catabolism.
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Regulation of genes in Streptomyces bacteria required for catabolism of lignin-derived aromatic compounds
Applied Microbiology and Biotechnology, 2010Co-Authors: Jennifer R. Davis, Jason K. SelloAbstract:The major utilization pathway for lignin-derived aromatic compounds in microorganisms is the β-ketoadipate pathway. Through this pathway, the aromatic compounds protocatechuate and catechol are converted to acetyl coenzyme A and succinyl coenzyme A. The enzymes of the protocatechuate branch of this pathway are encoded by the pca genes. Here, we describe a gene cluster in Streptomyces coelicolor containing the pca structural genes and a regulatory gene required for the catabolism of protocatechuate. We found that transcription of the structural genes in S. coelicolor is induced by protocatechuate and p -hydroxybenzoate. We also observed inducible transcription of pca structural genes in the ligninolytic strain Streptomyces Viridosporus ATCC 39115. Disruption of a gene encoding a putative MarR family transcription factor that is divergently transcribed from the pca structural genes resulted in constitutive transcription of the structural genes. Thus, the transcription factor encoded by this gene is an apparent negative regulator of pca gene transcription in S. coelicolor . Our findings suggest how Streptomyces bacteria could be engineered for and used in biotechnology for the utilization and degradation of lignin and lignin-derived aromatic compounds.
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Regulation of genes in Streptomyces bacteria required for catabolism of lignin-derived aromatic compounds
Applied microbiology and biotechnology, 2009Co-Authors: Jennifer R. Davis, Jason K. SelloAbstract:The major utilization pathway for lignin-derived aromatic compounds in microorganisms is the beta-ketoadipate pathway. Through this pathway, the aromatic compounds protocatechuate and catechol are converted to acetyl coenzyme A and succinyl coenzyme A. The enzymes of the protocatechuate branch of this pathway are encoded by the pca genes. Here, we describe a gene cluster in Streptomyces coelicolor containing the pca structural genes and a regulatory gene required for the catabolism of protocatechuate. We found that transcription of the structural genes in S. coelicolor is induced by protocatechuate and p-hydroxybenzoate. We also observed inducible transcription of pca structural genes in the ligninolytic strain Streptomyces Viridosporus ATCC 39115. Disruption of a gene encoding a putative MarR family transcription factor that is divergently transcribed from the pca structural genes resulted in constitutive transcription of the structural genes. Thus, the transcription factor encoded by this gene is an apparent negative regulator of pca gene transcription in S. coelicolor. Our findings suggest how Streptomyces bacteria could be engineered for and used in biotechnology for the utilization and degradation of lignin and lignin-derived aromatic compounds.
Jason K. Sello - One of the best experts on this subject based on the ideXlab platform.
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Regulation of genes in Streptomyces bacteria required for catabolism of lignin-derived aromatic compounds
Applied Microbiology and Biotechnology, 2010Co-Authors: Jennifer R. Davis, Jason K. SelloAbstract:The major utilization pathway for lignin-derived aromatic compounds in microorganisms is the β-ketoadipate pathway. Through this pathway, the aromatic compounds protocatechuate and catechol are converted to acetyl coenzyme A and succinyl coenzyme A. The enzymes of the protocatechuate branch of this pathway are encoded by the pca genes. Here, we describe a gene cluster in Streptomyces coelicolor containing the pca structural genes and a regulatory gene required for the catabolism of protocatechuate. We found that transcription of the structural genes in S. coelicolor is induced by protocatechuate and p -hydroxybenzoate. We also observed inducible transcription of pca structural genes in the ligninolytic strain Streptomyces Viridosporus ATCC 39115. Disruption of a gene encoding a putative MarR family transcription factor that is divergently transcribed from the pca structural genes resulted in constitutive transcription of the structural genes. Thus, the transcription factor encoded by this gene is an apparent negative regulator of pca gene transcription in S. coelicolor . Our findings suggest how Streptomyces bacteria could be engineered for and used in biotechnology for the utilization and degradation of lignin and lignin-derived aromatic compounds.
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Regulation of genes in Streptomyces bacteria required for catabolism of lignin-derived aromatic compounds
Applied microbiology and biotechnology, 2009Co-Authors: Jennifer R. Davis, Jason K. SelloAbstract:The major utilization pathway for lignin-derived aromatic compounds in microorganisms is the beta-ketoadipate pathway. Through this pathway, the aromatic compounds protocatechuate and catechol are converted to acetyl coenzyme A and succinyl coenzyme A. The enzymes of the protocatechuate branch of this pathway are encoded by the pca genes. Here, we describe a gene cluster in Streptomyces coelicolor containing the pca structural genes and a regulatory gene required for the catabolism of protocatechuate. We found that transcription of the structural genes in S. coelicolor is induced by protocatechuate and p-hydroxybenzoate. We also observed inducible transcription of pca structural genes in the ligninolytic strain Streptomyces Viridosporus ATCC 39115. Disruption of a gene encoding a putative MarR family transcription factor that is divergently transcribed from the pca structural genes resulted in constitutive transcription of the structural genes. Thus, the transcription factor encoded by this gene is an apparent negative regulator of pca gene transcription in S. coelicolor. Our findings suggest how Streptomyces bacteria could be engineered for and used in biotechnology for the utilization and degradation of lignin and lignin-derived aromatic compounds.