scholarly journals Wild type Bacillus subtilis treated with ethyl methyl sulphonate produced catabolite insensitive mutants with improved cellulase production

2021 ◽  
Author(s):  
Oladipo Olaniyi
Keyword(s):  
Bacillus Subtilis ◽  
Carbon Source ◽  
Enzyme Production ◽  
Cellulase Activity ◽  
Cellulase Production ◽  
Minimal Salt Medium ◽  
Salt Medium ◽  
Production Medium ◽  
Wild Type ◽  
Ethyl Methyl

Abstract The goal of this present investigation was to mutagenize Bacillus subtilis with Ethyl Methyl Sulphonate (EMS), screen the mutants for cellulase production and evaluate the influence of different glucose concentrations on their cellulase production potentials. The wild type B. subtilis was treated with 20, 40, 60 and 80 µl of EMS and the mutants generated were screened for cellulase production in minimal salt medium containing carboxylmethylcellulose (CMC) as the carbon source. Quantitatively, cellulase activity and protein contents were determined by dinitrosalicylic acid and Lowry methods respectively. Seven mutants were developed from each of the EMS concentration bringing the total to twenty-eight from all the concentrations. Approximately 14 and 57% of the mutants developed from 40 and 60µl of EMS had higher cellulase activities than the wild type, while none of the mutants developed from 20 and 80 µl of EMS had better activities than the wild type. The supplementation of 0.2, 0.5, 1.0 and 1.5% glucose in enzyme production medium caused approximately 100, 14, 29 and 14% cellulase repression respectively in the mutants developed from 60µl EMS. Mutants MSSS02 and MSSS05 were considered as catabolite insensitive mutants because their cellulase production were enhanced in comparison to wild type.

scholarly journals Comparative Analysis of Structural Variations Due to Genome Shuffling of Bacillus Subtilis VS15 for Improved Cellulase Production

2020 ◽  
Vol 21 (4) ◽  
pp. 1299 ◽  
Author(s):  
Soujanya Lakshmi Ega ◽  
Gene Drendel ◽  
Steve Petrovski ◽  
Eleonora Egidi ◽  
Ashley E. Franks ◽  
...  
Keyword(s):  
Bacillus Subtilis ◽  
Potassium Dichromate ◽  
Cellulase Activity ◽  
Genome Shuffling ◽  
Cellulase Production ◽  
Wild Type ◽  
Structural Variations ◽  
Renewable Biomass ◽  
Ethyl Methyl ◽  
A Genome

Cellulose is one of the most abundant and renewable biomass products used for the production of bioethanol. Cellulose can be efficiently hydrolyzed by Bacillus subtilis VS15, a strain isolate obtained from decomposing logs. A genome shuffling approach was implemented to improve the cellulase activity of Bacillus subtilis VS15. Mutant strains were created using ethyl methyl sulfonate (EMS), N-Methyl-N′ nitro-N-nitrosoguanidine (NTG), and ultraviolet light (UV) followed by recursive protoplast fusion. After two rounds of shuffling, the mutants Gb2, Gc8, and Gd7 were produced that had an increase in cellulase activity of 128%, 148%, and 167%, respectively, in comparison to the wild type VS15. The genetic diversity of the shuffled strain Gd7 and wild type VS15 was compared at whole genome level. Genomic-level comparisons identified a set of eight genes, consisting of cellulase and regulatory genes, of interest for further analyses. Various genes were identified with insertions and deletions that may be involved in improved celluase production in Gd7. Strain Gd7 maintained the capability of hydrolyzing wheatbran to glucose and converting glucose to ethanol by fermentation with Saccharomyces cerevisiae of the wild type VS17. This ability was further confirmed by the acidified potassium dichromate (K2Cr2O7) method.

scholarly journals Heterotrimeric G-Protein Signaling Is Required for Cellulose Degradation in Neurospora crassa

mBio ◽  
2020 ◽  
Vol 11 (6) ◽  
Author(s):  
Logan A. Collier ◽  
Arit Ghosh ◽  
Katherine A. Borkovich
Keyword(s):  
G Protein ◽  
Cellulose Degradation ◽  
Cellulase Activity ◽  
Cellulase Production ◽  
Camp Signaling ◽  
G Protein Signaling ◽  
Protein Signaling ◽  
Wild Type ◽  
Posttranscriptional Control ◽  
Heterotrimeric G Protein Signaling

ABSTRACT The filamentous fungus Neurospora crassa decomposes lignocellulosic biomass to generate soluble sugars as carbon sources. In this study, we investigated a role for heterotrimeric G-protein signaling in cellulose degradation. Loss of the Gα subunit genes gna-1 and gna-3, the Gβ subunit genes gnb-1 and cpc-2, the Gγ gene gng-1, or the gene for downstream effector adenylyl cyclase (cr-1) resulted in loss of detectable cellulase activity. This defect was also observed in strains expressing a constitutively active version of gna-3 (gna-3Q208L). We found that GNA-1 levels are greatly reduced in Δgna-3, Δgnb-1, and Δgng-1 strains, likely contributing to cellulase defects in these genetic backgrounds. The observation that gna-3Q208L Δgnb-1 strains exhibit cellulase activity, despite greatly reduced levels of GNA-1 protein, is consistent with positive control of cellulase production by GNA-3 that is manifested in the absence of gnb-1. Expression patterns for five cellulase genes showed that Δgna-1, Δgnb-1, and Δgna-3 mutants produce less cellulase mRNA than the wild type, consistent with transcriptional regulation. Δcpc-2 mutants had wild-type levels of cellulase transcripts, suggesting posttranscriptional control. In contrast, results for Δcr-1 mutants support both transcriptional and posttranscriptional control of cellulase activity by cAMP signaling. Cellulase activity defects in Δgna-3 mutants were fully remediated by cAMP supplementation, consistent with GNA-3 operating upstream of cAMP signaling. In contrast, cAMP addition only partially corrected cellulase activity defects in Δgna-1 and Δgnb-1 mutants, suggesting participation of GNA-1 and GNB-1 in additional cAMP-independent pathways that control cellulase activity. IMPORTANCE Filamentous fungi are critical for the recycling of plant litter in the biosphere by degrading lignocellulosic biomass into simpler compounds for metabolism. Both saprophytic and pathogenic fungi utilize plant cell wall-degrading enzymes to liberate carbon for metabolism. Several studies have demonstrated a role for cellulase enzymes during infection of economically relevant crops by fungal pathogens. Especially in developing countries, severe plant disease means loss of entire crops, sometimes leading to starvation. In this study, we demonstrate that G-protein signaling is a key component of cellulase production. Therefore, understanding the role of G-protein signaling in the regulation of the unique metabolism of cellulose by these organisms can inform innovations in strain engineering of industrially relevant species for biofuel production and in combatting food shortages caused by plant pathogens.

scholarly journals Inositol Catabolism, a Key Pathway in Sinorhizobium meliloti for Competitive Host Nodulation

2010 ◽  
Vol 76 (24) ◽  
pp. 7972-7980 ◽  
Author(s):  
Petra R. A. Kohler ◽  
Jasmine Y. Zheng ◽  
Elke Schoffers ◽  
Silvia Rossbach
Keyword(s):  
Bacillus Subtilis ◽  
Carbon Source ◽  
Sole Carbon Source ◽  
Sinorhizobium Meliloti ◽  
Mutational Analysis ◽  
Nodule Occupancy ◽  
Nitrogen Fixing ◽  
Wild Type ◽  
Catabolic Pathway ◽  
Inositol Dehydrogenase

ABSTRACT The nitrogen-fixing symbiont of alfalfa, Sinorhizobium meliloti, is able to use myo-inositol as the sole carbon source. Putative inositol catabolism genes (iolA and iolRCDEB) have been identified in the S. meliloti genome based on their similarities with the Bacillus subtilis iol genes. In this study, functional mutational analysis revealed that the iolA and iolCDEB genes are required for growth not only with the myo-isomer but also for growth with scyllo- and d-chiro-inositol as the sole carbon source. An additional, hypothetical dehydrogenase of the IdhA/MocA/GFO family encoded by the smc01163 gene was found to be essential for growth with scyllo-inositol, whereas the idhA-encoded myo-inositol dehydrogenase was responsible for the oxidation of d-chiro-inositol. The putative regulatory iolR gene, located upstream of iolCDEB, encodes a repressor of the iol genes, negatively regulating the activity of the myo- and the scyllo-inositol dehydrogenases. Mutants with insertions in the iolA, smc01163, and individual iolRCDE genes could not compete against the wild type in a nodule occupancy assay on alfalfa plants. Thus, a functional inositol catabolic pathway and its proper regulation are important nutritional or signaling factors in the S. meliloti-alfalfa symbiosis.

Isolation of endo-1,4-b-D-glucanase producing Bacillus subtilis sp. from fermented foods and enhanced enzyme production bydeveloping the mutant strain

2017 ◽  
Author(s):  
Jakyeom Seo ◽  
Eunhye Park ◽  
Sung Sill Leek ◽  
Byeongwoo Kim ◽  
Teaksoon Shin ◽  
...  
Keyword(s):  
Bacillus Subtilis ◽  
Mutant Strain ◽  
Enzyme Production ◽  
Methyl Cellulose ◽  
Feed Additives ◽  
Fermented Foods ◽  
Wild Type ◽  
Cellulolytic Bacteria ◽  
16S Rrna Analysis ◽  
Significant Difference

Cellulolytic bacteria living in food can be applied to microbial feed additives to improve fiber digestion in animal feeds. In this study, a cellulase-producing bacteria was isolated from salted clam and treated with physical or chemical agents to enhance their enzyme production. The bacteria was identified as a strain of Bacillus subtilis on the basis of 16S rRNA analysis. Endo-1,4-b-D-glucanase (endoglucanase) was produced by the wild type using 0.4% carboxy-methyl-cellulose as a carbon source with maximal activity (0.04 U/mL) after 24 h incubation. Insoluble cellulose and oat spelt xylan were also used as carbon sources for investigation of exoglucanase and xylanase, however, these enzymes were not found in the culture supernatant. Maximum endoglucanase activity of Bacillus subtilis sp. was measured at 50°C and pH 5, respectively. Then, the strain was subjected to classical mutagenesis (UV-irradiation and chemical treatment) to improve endoglucanase production. A mutant strain, P11 treated with ethyl methyl sulfonate was finally selected. Mutant P11 was sub-cultured and tested for endoglucanase production, which was 0.05 U/mL after 24 h growth. The significant difference of endoglucanase production between wild type and mutant P11 was prolonged to 10th generation. Thus, the mutant strain was found to have enhanced endoglucanase production.

scholarly journals Optimization for the Production of Cellulase Enzyme from Municipal Solid Waste Residue by Two Novel Cellulolytic Fungi

2011 ◽  
Vol 2011 ◽  
pp. 1-8 ◽  
Author(s):  
S. P. Gautam ◽  
P. S. Bundela ◽  
A. K. Pandey ◽  
Jamaluddin Khan ◽  
M. K. Awasthi ◽  
...  
Keyword(s):  
Municipal Solid Waste ◽  
Solid Waste ◽  
Enzyme Production ◽  
Nitrogen Sources ◽  
Cellulase Production ◽  
Production Medium ◽  
Optimum Temperature ◽  
Carbon And Nitrogen Sources ◽  
Carbon And Nitrogen ◽  
Alternative Carbon Source

The main purpose of this study is to reduce the production cost of cellulase by optimizing the production medium and using an alternative carbon source such as municipal solid waste residue. In the present investigation, we aim to isolate the two novel cellulase producing fungi (Aspergillus niger and Trichoderma sp.) from municipal solid waste. Municipal solid waste residue (4-5% (w/v)) and peptone and yeast extract (1.0% (w/v)) were found to be the best combination of carbon and nitrogen sources for the production of cellulase by A. niger and Trichoderma sp. Optimum temperature and pH of the medium for the cellulase production by A. niger were 40°C and 6-7, whereas those for the production of cellulase by Trichoderma sp. were 45°C and 6.5. Cellulase production from A. niger and Trichoderma sp. can be an advantage as the enzyme production rate is normally higher as compared to other fungi.

scholarly journals Process Design and Economics of On-Site Cellulase Production on Various Carbon Sources in a Softwood-Based Ethanol Plant

2010 ◽  
Vol 2010 ◽  
pp. 1-8 ◽  
Author(s):  
Zsolt Barta ◽  
Krisztina Kovacs ◽  
Kati Reczey ◽  
Guido Zacchi
Keyword(s):  
Carbon Source ◽  
Enzyme Production ◽  
Ethanol Yield ◽  
Liquid Fraction ◽  
Carbon Sources ◽  
Cellulase Production ◽  
Raw Material ◽  
Steam Pretreatment ◽  
Whole Process ◽  
Feasible Alternative

On-site cellulase enzyme fermentation in a softwood-to-ethanol process, based on SO2-catalysed steam pretreatment followed by simultaneous saccharification and fermentation, was investigated from a techno-economic aspect using Aspen Plus© and Aspen Icarus Process Evaluator© softwares. The effect of varying the carbon source of enzyme fermentation, at constant protein and mycelium yields, was monitored through the whole process. Enzyme production step decreased the overall ethanol yield (270 L/dry tonne of raw material in the case of purchased enzymes) by 5–16 L/tonne. Capital cost was found to be the main cost contributor to enzyme fermentation, constituting to 60–78% of the enzyme production cost, which was in the range of 0.42–0.53 SEK/L ethanol. The lowest minimum ethanol selling prices (4.71 and 4.82 SEK/L) were obtained in those scenarios, where pretreated liquid fraction supplemented with molasses was used as carbon source. In some scenarios, on-site enzyme fermentation was found to be a feasible alternative.

scholarly journals Optimization of Cellulase Production from Bacteria Isolated from Soil

2013 ◽  
Vol 2013 ◽  
pp. 1-7 ◽  
Author(s):  
Sonia Sethi ◽  
Aparna Datta ◽  
B. Lal Gupta ◽  
Saksham Gupta
Keyword(s):  
Bacillus Subtilis ◽  
Carbon Source ◽  
Pseudomonas Fluorescens ◽  
Carbon Sources ◽  
Nitrogen Sources ◽  
Fermentation Medium ◽  
Cellulase Production ◽  
Culture Conditions ◽  
Optimum Conditions ◽  
E Coli

Cellulase-producing bacteria were isolated from soil and identified as Pseudomonas fluorescens, Bacillus subtilIs, E. coli, and Serratia marcescens. Optimization of the fermentation medium for maximum cellulase production was carried out. The culture conditions like pH, temperature, carbon sources, and nitrogen sources were optimized. The optimum conditions found for cellulase production were 40°C at pH 10 with glucose as carbon source and ammonium sulphate as nitrogen source, and coconut cake stimulates the production of cellulase. Among bacteria, Pseudomonas fluorescens is the best cellulase producer among the four followed by Bacillus subtilis, E. coli, and Serratia marscens.

scholarly journals Identification of a Pathogenicity Locus, rpfA, in Erwinia carotovora subsp. carotovora That Encodes a Two-Component Sensor-Regulator Protein

1997 ◽  
Vol 10 (3) ◽  
pp. 407-415 ◽  
Author(s):  
Reid D. Frederick ◽  
Jiliang Chiu ◽  
Jeffrey L. Bennetzen ◽  
Autar K. Handa
Keyword(s):  
Xanthomonas Campestris ◽  
Genetic Locus ◽  
Cellulase Activity ◽  
Pectate Lyase ◽  
Erwinia Carotovora ◽  
Cellulase Production ◽  
Extracellular Protease ◽  
Wild Type ◽  
Reading Frame ◽  
Two Component

A mutant of Erwinia carotovora subsp. carotovora, AH2552, created by a Mud1 insertion was found to be reduced in plant pathogenicity and deficient in extracellular protease and cellulase activity, although it produced normal levels of pectate lyase and polygalacturonase. A cosmid clone, pEC462, was isolated from a wild-type E. carotovora subsp. carotovora DNA library that concomitantly restored pathogenicity and protease and cellulase activities of AH2552 to wild-type levels when present in trans. The genetic locus that was disrupted in AH2552 by insertion of Mud1 has been designated rpfA, for regulator of pathogenicity factors. Sequencing of the rpfA region identified an open reading frame of 2,787 bp, and the predicted 929-amino acid polypeptide shared high identity with several two-component sensor-regulator proteins: BarA from Escherichia coli, ApdA from Pseudomonas fluorescens, PheN from P. tolaasii, RepA from P. viridiflava, LemA from P. syringae pv. syringae, and RpfC from Xanthomonas campestris pv. campestris. The RpfA locus described in this study encodes a putative sensor kinase protein that is involved in both extracellular protease and cellulase production and the pathogenicity of E. carotovora subsp. carotovora on potato tubers.

scholarly journals Induction of Cellulase Biosynthesis by Cellobiose Octaacetate in Aspergillus humicola

1970 ◽  
Vol 23 (2) ◽  
pp. 174-176 ◽  
Author(s):  
Meher Nigad Nipa ◽  
Sharmin Sultana ◽  
M Abdul Hakim
Keyword(s):  
Carbon Source ◽  
Enzyme Production ◽  
Maximum Activity ◽  
Production Medium ◽  
Cmcase Activity ◽  
Initial Ph ◽  
Enzyme Biosynthesis

Aspergillus humicola, one of the major cellulase-producing fungi, was used in this study for carboxymethylcellulase (CMCase) production using Winstead's basal broth supplanted with cellobiose octaacetate (COA), a synthetic carbon source. Under all conditions, the enzyme biosynthesis was remarkably increased when the inducer COA was added to the production medium containing carboxymethylcellulose (CMC). Maximum enzyme production (1.62 U/ml) was achieved in COA-containing at 37°C. The enzyme production was highest at initial pH 5.5 and after 7 days incubation. The enzyme exhibited maximum activity at 40°C with a reaction pH 5.5. CMCase activity was inhibited by its own substrate CMC at concentration higher than 1.0%. The study clearly demonstrated that COA is a good inducer for extracellular CMCase production by the fungus. Keywords: Aspergillus humicola, Carboxymethylcellulase (CMCase), Carboxymethylcellulose (CMC), Cellobiose octaacetate (COA)DOI: http://dx.doi.org/10.3329/bjm.v23i2.889 Bangladesh J Microbiol, Volume 23, Number 2, December 2006, pp 174-176

scholarly journals Probing cellulolytic yeast from forest ecosystem for the saccharification of Napier fodder biomass

2021 ◽  
Vol 42 (4(SI)) ◽  
pp. 1152-1161
Author(s):  
M.G. Valliammai ◽  
◽  
N.O. Gopal ◽  
R. Anandham ◽  
◽  
...  
Keyword(s):  
Carbon Source ◽  
Statistical Model ◽  
Forest Ecosystem ◽  
Cellulase Activity ◽  
Cellulase Production ◽  
Media Composition ◽  
Yeast Isolate ◽  
Trichosporon Asahii ◽  
Fuels And Chemicals ◽  
The Media

Aim: This study aimed to search for novel cellulolytic isolates with high cellulase titre for the production of fuels and chemicals. Methodology: The yeast isolate YES5 isolated from the forest soil was screened for cellulase production. The cellulase activity of YES5 was optimized via RSM. The saccharification potential of YES5 using Napier biomass as substrate was evalauted. Results: The maximum cellulase activity obtained after optimizing pH, temperature, and incubation period was 35.70 U. A reliable statistical model was developed for maximizing the cellulase activity in YES5 Trichosporon asahii. The cellulase activity was 23.87U, when carbon source in CMC medium was replaced by Napier biomass. The maximum saccharification potential of 33.15% was observed on 3rd day. Interpretation: The study of optimizing the media composition of Trichosporon asahii cellulase using Napier biomass, a natural source of carbon for maximizing the cellulase production via RSM, is first of its kind.

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