scholarly journals Ralstonia Eutropha and the Production of Value Added Products: Metabolic Background of the Wild-Type Strain and its Role as a Diverse, Genetically-Engineered Biocatalyst Organism

2016 ◽  
pp. 265-347 ◽  
Keyword(s):  
Type Strain ◽  
Wild Type Strain ◽  
Ralstonia Eutropha ◽  
Value Added ◽  
Genetically Engineered ◽  
Wild Type ◽  
Value Added Products

scholarly journals Assessing the Potential of Newly Isolated Pichia Fermentans for Xylitol Production Using Non-Detoxified Xylose Rich Pre-Hydrolysate Derived from Sugarcane Bagasse

2020 ◽  
Author(s):  
Ashish A Prabhu ◽  
Ekkarin Bosakornranut ◽  
Yassin Amraoui ◽  
Deepti Agarwal ◽  
Frederic Coulon ◽  
...  
Keyword(s):  
Sugarcane Bagasse ◽  
Type Strain ◽  
Shake Flask ◽  
Wild Type Strain ◽  
Xylose Reductase ◽  
Value Added ◽  
Xylitol Production ◽  
Wild Type ◽  
Conversion Yield ◽  
Cell Factories

Abstract Background: Integrated management of hemicellulosic fraction and its economical transformation to value-added products is the key driver towards sustainable second-generation biorefineries. In this aspect microbial cell factories are harnessed for sustainable production of biochemicals by valorising C5 and C6 sugars generated from agro-industrial waste. However, most of the strains can effectively consume C6 sugars but lacks pentose metabolism pathway. The effective utilization of both pentose and hexose sugars is key for economical biorefinery. Results: In the current study, the ability of a newly isolated xylose assimilating Pichia fermentans was explored for xylitol production. The wild type strain robustly grew on xylose and produced xylitol with >40% conversion yield. Mutagenesis with ethyl methanesulphonate (EMS) yielded seven mutants. The mutant obtained after 15 min exposure, exhibited best xylose bioconversion efficiency. This mutant under shake flask conditions produced maximum xylitol titre and yield of 34.0 g/L and 0.68 g/g, respectively. oweverHoHHHoHowever, under same conditions, the control wild type strain accumulated 27.0 g/L xylitol with a conversion yield of 0.45 g/g. Improved performance of the mutant was attributed to 34.6% activity enhancement in xylose reductase with simultaneous reduction of xylitol dehydrogenase activity by 22.9%. Later, the culture medium was optimized using statistical design and validated at shake flask and bioreactor level. Bioreactor studies affirmed the competence of mutant in xylitol accumulation. The xylitol titre and yield obtained with pure xylose were 98.9 g/L and 0.67 g/g, respectively while xylitol produced using non-detoxified xylose rich pre-hydrolysate from sugarcane bagasse was 79.0 g/L with an overall yield of 0.54 g/g. Conclusion: This study established the potential of P. fermentans in successfully valorising the hemicellulosic fraction for sustainable xylitol production.

scholarly journals Importance of Different tfd Genes for Degradation of Chloroaromatics by Ralstonia eutropha JMP134

2002 ◽  
Vol 184 (15) ◽  
pp. 4054-4064 ◽  
Author(s):  
Iris Plumeier ◽  
Danilo Pérez-Pantoja ◽  
Sabina Heim ◽  
Bernardo González ◽  
Dietmar H. Pieper
Keyword(s):  
Type Strain ◽  
Wild Type Strain ◽  
Ralstonia Eutropha ◽  
Major Product ◽  
Kinetic Properties ◽  
Gene Module ◽  
Wild Type ◽  
In The Wild ◽  
Gene Modules ◽  
Chloromuconate Cycloisomerase

ABSTRACT The tfdC I D I E I F I, and tfdD II C II E II F II gene modules of plasmid pJP4 of Ralstonia eutropha JMP134 encode complete sets of functional enzymes for the transformation of chlorocatechols into 3-oxoadipate, which are all expressed during growth on 2,4-dichlorophenoxyacetate (2,4-D). However, activity of tfd I-encoded enzymes was usually higher than that of tfd II-encoded enzymes, both in the wild-type strain grown on 2,4-D and in 3-chlorobenzoate-grown derivatives harboring only one tfd gene module. The tfdD II-encoded chloromuconate cycloisomerase exhibited special kinetic properties, with high activity against 3-chloromuconate and poor activity against 2-chloromuconate and unsubstituted muconate, thus explaining the different phenotypic behaviors of R. eutropha strains containing different tfd gene modules. The enzyme catalyzes the formation of an equilibrium between 2-chloromuconate and 5-chloro- and 2-chloromuconolactone and very inefficiently catalyzes dehalogenation to form trans-dienelactone as the major product, thus differing from all (chloro)muconate cycloisomerases described thus far.

Polyhydroxyalkanoate production in wild-type and engineered Ralstonia eutropha: carbon flow from central metabolism to storage for value added products

2012 ◽  
Vol 29 ◽  
pp. S9-S10
Author(s):  
Christopher Brigham ◽  
Sebastian Riedel ◽  
Jingnan Lu ◽  
Matthew Neal ◽  
John W. Quimby ◽  
...  
Keyword(s):  
Ralstonia Eutropha ◽  
Value Added ◽  
Carbon Flow ◽  
Central Metabolism ◽  
Wild Type ◽  
Value Added Products

scholarly journals Molecular Monitoring of Wild-Type and Genetically Engineered Colletotrichum coccodes Biocontrol Strains In Planta

Plant Disease ◽  
2006 ◽  
Vol 90 (12) ◽  
pp. 1504-1510 ◽  
Author(s):  
A. L. Dauch ◽  
B. Ahn ◽  
A. K. Watson ◽  
P. Seguin ◽  
S. H. Jabaji-Hare
Keyword(s):  
Type Strain ◽  
Wild Type Strain ◽  
Quantitative Polymerase Chain Reaction ◽  
Fungal Growth ◽  
Genetically Engineered ◽  
Colletotrichum Coccodes ◽  
Wild Type ◽  
In Planta ◽  
Molecular Monitoring ◽  
Dna Amounts

Two strains of Colletotrichum coccodes, the wild type (DAOM 183088) and T-20a, engineered with the necrosis- and ethylene-inducing peptide (NEP1) gene for hypervirulence on velvetleaf (Abutilon theophrasti, Medik.), were monitored in planta for the first 2 weeks after infection. Real-time quantitative polymerase chain reaction (QPCR) was used to assess the extent of colonization of both strains on velvetleaf using SYBR Green chemistry. Quantification of both strains was successful as soon as the conidia were sprayed on the leaves and up to 14 days after infection. The increase in fungal DNA amounts corroborated with the appearance of necrotic lesions on velvetleaf leaves infected with the wild-type strain. The wild-type C. coccodes was more efficient at infecting velvetleaf than the transgenic T-20a strain. In addition, detection of host DNA allowed us to quantitatively monitor the decrease in plant DNA amounts in response to wild-type strain infection. Expression of the NEP1 transgene by conventional retro-transcription (RT)-PCR was absent from T-20a growing on either V8 agar or in planta, suggesting that the gene may be silenced. The application of QPCR to monitor fungal growth was proven to detect the target organisms in planta prior to the appearance of symptoms.

scholarly journals Enhanced xylitol production using non-detoxified xylose rich pre-hydrolysate from sugarcane bagasse by newly isolated Pichia fermentans

2020 ◽  
Vol 13 (1) ◽  
Author(s):  
Ashish A. Prabhu ◽  
Ekkarin Bosakornranut ◽  
Yassin Amraoui ◽  
Deepti Agrawal ◽  
Frederic Coulon ◽  
...  
Keyword(s):  
Sugarcane Bagasse ◽  
Type Strain ◽  
Shake Flask ◽  
Wild Type Strain ◽  
Value Added ◽  
Chemical Mutagenesis ◽  
Xylitol Production ◽  
Wild Type ◽  
Conversion Yield ◽  
Cell Factories

Abstract Background Integrated management of hemicellulosic fraction and its economical transformation to value-added products is the key driver towards sustainable lignocellulosic biorefineries. In this aspect, microbial cell factories are harnessed for the sustainable production of commercially viable biochemicals by valorising C5 and C6 sugars generated from agro-industrial waste. However, in the terrestrial ecosystem, microbial systems can efficiently consume glucose. On the contrary, pentose sugars are less preferred carbon source as most of the microbes lack metabolic pathway for their utilization. The effective utilization of both pentose and hexose sugars is key for economical biorefinery. Results Bioprospecting the food waste and selective enrichment on xylose-rich medium led to screening and isolation of yeast which was phylogenetically identified as Pichia fermentans. The newly isolated xylose assimilating yeast was explored for xylitol production. The wild type strain robustly grew on xylose and produced xylitol with > 40% conversion yield. Chemical mutagenesis of isolated yeast with ethyl methanesulphonate (EMS) yielded seven mutants. The mutant obtained after 15 min EMS exposure, exhibited best xylose bioconversion efficiency. This mutant under shake flask conditions produced maximum xylitol titer and yield of 34.0 g/L and 0.68 g/g, respectively. However, under the same conditions, the control wild type strain accumulated 27.0 g/L xylitol with a conversion yield of 0.45 g/g. Improved performance of the mutant was attributed to 34.6% activity enhancement in xylose reductase with simultaneous reduction of xylitol dehydrogenase activity by 22.9%. Later, the culture medium was optimized using statistical design and validated at shake flask and bioreactor level. Bioreactor studies affirmed the competence of the mutant for xylitol accumulation. The xylitol titer and yield obtained with pure xylose were 98.9 g/L and 0.67 g/g, respectively. In comparison, xylitol produced using non-detoxified xylose rich pre-hydrolysate from sugarcane bagasse was 79.0 g/L with an overall yield of 0.54 g/g. Conclusion This study demonstrates the potential of newly isolated P. fermentans in successfully valorising the hemicellulosic fraction for the sustainable xylitol production.

scholarly journals Carbonic Anhydrase Is Essential for Growth of Ralstonia eutropha at Ambient CO2 Concentrations

2002 ◽  
Vol 184 (18) ◽  
pp. 5018-5026 ◽  
Author(s):  
Bernhard Kusian ◽  
Dieter Sültemeyer ◽  
Botho Bowien
Keyword(s):  
Carbonic Anhydrase ◽  
Type Strain ◽  
Wild Type Strain ◽  
Ralstonia Eutropha ◽  
Maximal Activity ◽  
Metabolic Function ◽  
Wild Type ◽  
Air Concentration ◽  
Ambient Co2

ABSTRACT Mutant strain 25-1 of the facultative chemoautotroph Ralstonia eutropha H16 had previously been shown to exhibit an obligately high-CO2-requiring (HCR) phenotype. Although the requirement varied with the carbon and energy sources utilized, none of these conditions allowed growth at the air concentration of CO2. In the present study, a gene designated can and encoding a β-carbonic anhydrase (CA) was identified as the site altered in strain 25-1. The mutation caused a replacement of the highly conserved glycine residue 98 by aspartate in Can. A can deletion introduced into wild-type strain H16 generated mutant HB1, which showed the same HCR phenotype as mutant 25-1. Overexpression of can in Escherichia coli and mass spectrometric determination of CA activity demonstrated that can encodes a functional CA. The enzyme is inhibited by ethoxyzolamide and requires 40 mM MgSO4 for maximal activity. Low but significant CA activities were detected in wild-type H16 but not in mutant HB1, strongly suggesting that the CA activity of Can is essential for growth of the wild type in the presence of low CO2 concentrations. The HCR phenotype of HB1 was overcome by complementation with heterologous CA genes, indicating that growth of the organism at low CO2 concentrations requires sufficient CA activity rather than the specific function of Can. The metabolic function(s) depending on CA activity remains to be identified.

scholarly journals Enhanced xylitol production using non-detoxified xylose rich pre-hydrolysate from sugarcane bagasse by newly isolated Pichia fermentans

2020 ◽  
Author(s):  
Ashish A Prabhu ◽  
Ekkarin Bosakornranut ◽  
Yassin Amraoui ◽  
Deepti Agarwal ◽  
Frederic Coulon ◽  
...  
Keyword(s):  
Sugarcane Bagasse ◽  
Type Strain ◽  
Shake Flask ◽  
Wild Type Strain ◽  
Value Added ◽  
Chemical Mutagenesis ◽  
Xylitol Production ◽  
Wild Type ◽  
Conversion Yield ◽  
Isolation And Purification

Abstract Background: Integrated management of hemicellulosic fraction and its economical transformation to value-added products is the key driver towards sustainable lignocellulosic biorefineries. In this aspect, microbial cell factories are harnessed for the sustainable production of commercially viable biochemicals by valorising C5 and C6 sugars generated from agro-industrial waste. However, in the terrestrial ecosystem, microbial systems can efficiently consume glucose. On the contrary, pentose sugars are less preferred carbon source as most of the microbes lack metabolic pathway for their utilization. The effective utilization of both pentose and hexose sugars is key for economical biorefinery.Results: Bioprospecting the food waste and selective enrichment on xylose-rich medium led to isolation and purification of yeast which was phylogenetically identified as Pichia fermentans. The newly isolated xylose assimilating yeast was explored for xylitol production. The wild type strain robustly grew on xylose and produced xylitol with >40% conversion yield. Chemical mutagenesis of isolated yeast with ethyl methanesulphonate (EMS) yielded seven mutants. The mutant obtained after 15 min exposure, exhibited best xylose bioconversion efficiency. This mutant under shake flask conditions produced maximum xylitol titer and yield of 34.0 g/L and 0.68 g/g, respectively. However, under same conditions, the control wild type strain accumulated 27.0 g/L xylitol with a conversion yield of 0.45 g/g. Improved performance of the mutant was attributed to 34.6% activity enhancement in xylose reductase with simultaneous reduction of xylitol dehydrogenase activity by 22.9%. Later, the culture medium was optimized using statistical design and validated at shake flask and bioreactor level. Bioreactor studies affirmed the competence of mutant for xylitol accumulation. The xylitol titer and yield obtained with pure xylose were 98.9 g/L and 0.67 g/g, respectively. In comparison, xylitol produced using non-detoxified xylose rich pre-hydrolysate from sugarcane bagasse was 79.0 g/L with an overall yield of 0.54 g/g.Conclusion: This study demonstrates the potential of newly isolated P. fermentans in successfully valorising the hemicellulosic fraction for sustainable xylitol production.

scholarly journals Pathogenicity ofTreponema denticolaWild-Type and Mutant Strain Tested by an Active Mode of Periodontal Infection Using Microinjection

2012 ◽  
Vol 2012 ◽  
pp. 1-4 ◽  
Author(s):  
Jacques Izard ◽  
Hajime Sasaki ◽  
Ralph Kent
Keyword(s):  
Mutant Strain ◽  
Type Strain ◽  
Wild Type Strain ◽  
Genetically Engineered ◽  
Bacterial Attachment ◽  
Lesion Site ◽  
Wild Type ◽  
Passive Mode ◽  
Active Mode ◽  
Periodontal Infection

The available passive mode of periodontal infections in mice requires high efficiency of bacterial attachment and invasiveness and is not always suitable to test the pathogenicity of genetically engineered mutant strains. We developed an active mode of oral infection, using microinjection in the marginal gingiva of mice, to test the pathogenicity of a genetically engineeredTreponema denticolamutant strain deficient in intermediate-like filaments, compared to the wild-type strain. This targeted mode of infection inoculates the bacterial strain to be tested directly at a lesion site (needle entry point) located at the future periodontal lesion site. The efficiency ofT. denticolawild-type strain to elicit bone loss contrasted with the lack of pathogenicity of the intermediate-like filament deficient mutant strain in comparison to the sham infection. The periodontal microinjection oral model in mice can be used for a variety of applications complementary to the passive mode of periodontal infection in context of pathogenicity testing.

scholarly journals Enhanced xylitol production using non-detoxified xylose rich pre-hydrolysate from sugarcane bagasse by newly isolated Pichia fermentans

2020 ◽  
Author(s):  
Ashish A Prabhu ◽  
Ekkarin Bosakornranut ◽  
Yassin Amraoui ◽  
Deepti Agarwal ◽  
Frederic Coulon ◽  
...  
Keyword(s):  
Sugarcane Bagasse ◽  
Type Strain ◽  
Shake Flask ◽  
Wild Type Strain ◽  
Value Added ◽  
Chemical Mutagenesis ◽  
Xylitol Production ◽  
Wild Type ◽  
Conversion Yield ◽  
Isolation And Purification

Abstract Background: Integrated management of hemicellulosic fraction and its economical transformation to value-added products is the key driver towards sustainable lignocellulosic biorefineries. In this aspect, microbial cell factories are harnessed for the sustainable production of commercially viable biochemicals by valorising C5 and C6 sugars generated from agro-industrial waste. However, in the terrestrial ecosystem, microbial systems can efficiently consume glucose. On the contrary, pentose sugars are less preferred carbon source as most of the microbes lack metabolic pathway for their utilization. The effective utilization of both pentose and hexose sugars is key for economical biorefinery. Results: Bioprospecting the food waste and selective enrichment on xylose-rich medium led to isolation and purification of yeast which was phylogenetically identified as Pichia fermentans. The newly isolated xylose assimilating yeast was explored for xylitol production. The wild type strain robustly grew on xylose and produced xylitol with >40% conversion yield. Chemical mutagenesis of isolated yeast with ethyl methanesulphonate (EMS) yielded seven mutants. The mutant obtained after 15 min exposure, exhibited best xylose bioconversion efficiency. This mutant under shake flask conditions produced maximum xylitol titer and yield of 34.0 g/L and 0.68 g/g, respectively. oweverHoHHHoHowever, under same conditions, the control wild type strain accumulated 27.0 g/L xylitol with a conversion yield of 0.45 g/g. Improved performance of the mutant was attributed to 34.6% activity enhancement in xylose reductase with simultaneous reduction of xylitol dehydrogenase activity by 22.9%. Later, the culture medium was optimized using statistical design and validated at shake flask and bioreactor level. Bioreactor studies affirmed the competence of mutant for xylitol accumulation. The xylitol titer and yield obtained with pure xylose were 98.9 g/L and 0.67 g/g, respectively. In comparison, xylitol produced using non-detoxified xylose rich pre-hydrolysate from sugarcane bagasse was 79.0 g/L with an overall yield of 0.54 g/g. Conclusion: This study demonstrates the potential of newly isolated P. fermentans in successfully valorising the hemicellulosic fraction for sustainable xylitol production.

scholarly journals Effects of Mutations of RAD50, RAD51, RAD52, and Related Genes on Illegitimate Recombination in Saccharomyces cerevisiae

Genetics ◽  
1996 ◽  
Vol 142 (2) ◽  
pp. 383-391 ◽  
Author(s):  
Yasumasa Tsukamoto ◽  
Jun-ichi Kato ◽  
Hideo Ikeda
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Quantitative Analysis ◽  
Structural Analysis ◽  
Recombination Rate ◽  
Type Strain ◽  
Negative Selection ◽  
Wild Type Strain ◽  
Illegitimate Recombination ◽  
Wild Type ◽  
In The Wild

Abstract To examine the mechanism of illegitimate recombination in Saccharomyces cerevisiae, we have developed a plasmid system for quantitative analysis of deletion formation. A can1 cyh2 cell carrying two negative selection markers, the CAN1 and CYH2 genes, on a YCp plasmid is sensitive to canavanine and cycloheximide, but the cell becomes resistant to both drugs when the plasmid has a deletion over the CAN1 and CYH2 genes. Structural analysis of the recombinant plasmids obtained from the resistant cells showed that the plasmids had deletions at various sites of the CAN1-CYH2 region and there were only short regions of homology (1-5 bp) at the recombination junctions. The results indicated that the deletion detected in this system were formed by illegitimate recombination. Study on the effect of several rad mutations showed that the recombination rate was reduced by 30-, 10-, 10-, and 10-fold in the rad52, rad50, mre11, and xrs2 mutants, respectively, while in the rud51, 54, 55, and 57 mutants, the rate was comparable to that in the wild-type strain. The rad52 mutation did not affect length of homology at junction sites of illegitimate recombination.

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