scholarly journals Multilevel optimisation of anaerobic ethyl acetate production in engineered Escherichia coli

2020 ◽  
Author(s):  
Anna Christina Bohnenkamp ◽  
Aleksander J. Kruis ◽  
Astrid E. Mars ◽  
Rene H. Wijffels ◽  
John van der Oost ◽  
...  
Keyword(s):  
Ethyl Acetate ◽  
Production Efficiency ◽  
Yeast Species ◽  
Coli Strain ◽  
E Coli ◽  
Alcohol Acetyltransferase ◽  
Enhanced Production ◽  
Key Enzyme ◽  
Encoding Genes ◽  
Multilevel Optimisation

Abstract Background Ethyl acetate is a widely used industrial solvent that is currently produced by chemical conversions from fossil resources. Several yeast species are able to convert sugars to ethyl acetate under aerobic conditions. However, performing ethyl acetate synthesis anaerobically may result in enhanced production efficiency, making the process economically more viable. Results We engineered an E. coli strain that is able to convert glucose to ethyl acetate as the main fermentation product under anaerobic conditions. The key enzyme of the pathway is an alcohol acetyltransferase (AAT) that catalyses the formation of ethyl acetate from acetyl-CoA and ethanol. To select a suitable AAT, the ethyl acetate-forming capacities of Atf1 from Saccharomyces cerevisiae, Eat1 from Kluyveromyces marxianus and Eat1 from Wickerhamomyces anomalus were compared. Heterologous expression of the AAT-encoding genes under control of the inducible LacI/ T7 and XylS/ Pm promoters allowed optimisation of their expression levels. Conclusion Engineering efforts on protein and fermentation level resulted in an E. coli strain that anaerobically produced ethyl acetate from glucose at an unprecedented level, i.e. 0.48 C-mol/C-mol or 72% of the maximum pathway yield.

scholarly journals Multilevel optimisation of anaerobic ethyl acetate production in engineered Escherichia coli

2020 ◽  
Author(s):  
Anna Christina Bohnenkamp ◽  
Aleksander J. Kruis ◽  
Astrid E. Mars ◽  
Rene H. Wijffels ◽  
John van der Oost ◽  
...  
Keyword(s):  
Ethyl Acetate ◽  
Production Efficiency ◽  
Yeast Species ◽  
Anaerobic Conditions ◽  
E Coli ◽  
Alcohol Acetyltransferase ◽  
Enhanced Production ◽  
Key Enzyme ◽  
Encoding Genes ◽  
Multilevel Optimisation

Abstract BackgroundEthyl acetate is a widely used industrial solvent that is currently produced by chemical conversions from fossil resources. Several yeast species are able to convert sugars to ethyl acetate under aerobic conditions. However, performing ethyl acetate synthesis anaerobically may result in enhanced production efficiency, making the process economically more viable. ResultsWe engineered an E. coli strain that is able to convert glucose to ethyl acetate as the main fermentation product under anaerobic conditions. The key enzyme of the pathway is an alcohol acetyltransferase (AAT) that catalyses the formation of ethyl acetate from acetyl-CoA and ethanol. To select a suitable AAT, the ethyl acetate-forming capacities of Atf1 from Saccharomyces cerevisiae, Eat1 from Kluyveromyces marxianus and Eat1 from Wickerhamomyces anomalus were compared. Heterologous expression of the AAT-encoding genes under control of the inducible LacI/T7 and XylS/Pm promoters allowed optimisation of their expression levels. ConclusionEngineering efforts on protein and fermentation level resulted in an E. coli strain that anaerobically produced ethyl acetate from glucose at an unprecedented level, i.e. 0.48 C-mol/C-mol or 72% of the maximum pathway yield.

scholarly journals High cell density cultivation of a recombinant E. coli strain expressing a key enzyme in bioengineered heparin production

2013 ◽  
Vol 97 (9) ◽  
pp. 3893-3900 ◽  
Author(s):  
Odile Francesca Restaino ◽  
Ujjwal Bhaskar ◽  
Priscilla Paul ◽  
Lingyun Li ◽  
Mario De Rosa ◽  
...  
Keyword(s):  
Cell Density ◽  
High Cell Density ◽  
Coli Strain ◽  
High Cell ◽  
High Cell Density Cultivation ◽  
E Coli ◽  
Bioengineered Heparin ◽  
Key Enzyme

scholarly journals Enhanced Production of Carboxymethylcellulase by Recombinant Escherichia coli Strain from Rice Bran with Shifts in Optimal Conditions of Aeration Rate and Agitation Speed on a Pilot-Scale

2019 ◽  
Vol 9 (19) ◽  
pp. 4083
Author(s):  
Chung-Il Park ◽  
Jae-Hong Lee ◽  
Jianhong Li ◽  
Jin-Woo Lee
Keyword(s):  
Escherichia Coli ◽  
Cell Growth ◽  
Pilot Scale ◽  
Aeration Rate ◽  
Coli Strain ◽  
Agitation Speed ◽  
Recombinant Escherichia Coli ◽  
Optimal Conditions ◽  
E Coli ◽  
Enhanced Production

The optimal conditions including the aeration rate and agitation speed of bioreactors for the production of carboxymethylcellulase (CMCase) by a recombinant Escherichia coli KACC 91335P, expressing CMCase gene of B. velezensis A-68, were different from those for its cell growth. The enhanced production of CMCase by E. coli KACC 91335P with the conventional multistage process needs at least two bioreactors. Shifts in the optimal conditions of the aeration rate and agitation speed of the bioreactor from the cell growth of E. coli KACC 91335P to those for its production of CMCase were investigated for development of the simple and economic process with the high productivity and low cost. The production of CMCase by E. coli KACC 91335P with shifts in the optimal conditions of the aeration rate and agitation speed from the cell growth to its production of CMCase in a 100 L pilot-scale bioreactor was 1.36 times higher than that with a fixed optimal conditions of the aeration rate and agitation speed for the production of CMCase and it was even 1.54 times higher than that with a fixed optimal conditions of the aeration rate and agitation speed for cell growth. The best time for the shift in the optimal conditions was found to be the mid-log phase of cell growth. Owing to the mixed-growth-associated production of CMCase by E. coli KACC 91335P, shifts in the optimal conditions of the aeration rate and agitation speed of bioreactors from the cell growth to its production of CMCase seemed to result in relatively more cells for the participation in its production of CMCase, which in turn enhanced its production of CMCase. The process with a simple control for shifts in the aeration rate and agitation speed of a bioreactor for the enhanced production of CMCase by E. coli KACC 91335P on the pilot-scale can be directly applied to the industrial-scaled production of cellulase.

scholarly journals Recombinant Escherichia coli Strain with Enhanced Production of Vibrio cholerae Neuraminidase

2019 ◽  
pp. 87-92
Author(s):  
E. V. Monakhova ◽  
G. V. Demidova ◽  
R. V. Pisanov ◽  
O. V. Duvanova ◽  
B. N. Mishan’kin
Keyword(s):  
Vibrio Cholerae ◽  
Pharmaceutical Preparations ◽  
Plasmid Vector ◽  
Coli Strain ◽  
Total Cell ◽  
Iptg Induction ◽  
E Coli ◽  
Neuraminidase Activity ◽  
Enhanced Production ◽  
Cloned Gene

Objectiveof this work was cloning of the Vibrio cholerae nanH gene as part of a plasmid vector, providing expression of foreign genes under the control of T5 promoter, and construction of a E. coli strain – producer of V. cholerae recombinant neuraminidase.Materials and methods. V. cholerae о1 strain served as a DNA donor, pQE30 – as a vector plasmid. The gene was PCR-amplifi ed, the cloning was carried out by means of conventional methods, performance of recombinants and localization of the required protein was determined based on the results of electrophoresis of cell lysates. Neuraminidase activity was identifi ed by fl uorescence in ultraviolet light after incubation with specifi c substrate (4-methylumbelliferyl-N-acetylneuraminic acid).Results and discussion. Recombinant plasmid pNanH, containing the cloned gene nanH V. cholerae, has been constructed. The gene is inserted into BamHI-PstI sites of the polylinker of pQE30. Expression of the cloned gene in the producer strain E. coli JM103pNanH occurs under the control of T5 promoter after isopropyl-1-thio-β-D-galactopyranoside (IPTG) induction. The strain shows neuraminidase activity. The recombinant NanH protein is accumulated in the producer’s cells in two forms. The fi rst form, with molecular mass (MM) of 89.5 kDa, is an unprocessed protein with the hexahistidine block (6His-tag) at its N-terminus, it is located in inclusion bodies. Its percentage is 5.6–6.6 % of the total cell proteins. The second one, with MM of 83 kDa, is found in the periplasmic space and corresponds to the mature NanH, its percentage being 3.4–3.8 %. The total percentage of both forms is 9–10 % of total cell proteins, which allows to consider the strain E. coli JM103pNanH to be a super-producer of the required protein. The strain may be used for purifi cation of NanH preparations for construction of specifi c diagnostic, therapeutic and pharmaceutical preparations as well as for investigation of the protein as a virulence/persistence factor of the pathogen. 

Transport of escherichia coli through soil to groundwater traced by randomly amplified polymorphic DNA (RAPD)

1997 ◽  
Vol 35 (11-12) ◽  
pp. 351-357 ◽  
Author(s):  
R. Rothmaier ◽  
A. Weidenmann ◽  
K. Botzenhart
Keyword(s):  
Negative Impact ◽  
Random Amplified Polymorphic Dna ◽  
Coli Strain ◽  
Soil Samples ◽  
Test Field ◽  
Liquid Manure ◽  
E Coli ◽  
Rapd Pattern ◽  
Geological Situation ◽  
Different Strains

Isolates (50) of E. coli obtained from liquid manure (20 bovine, 20 porcine) were genotyped using random amplified polymorphic DNA (RAPD). Typing revealed 9 and 14 different strains in bovine and porcine liquid manure respectively with no strains in common. One porcine strain, showing a simple RAPD pattern, was subcultured and spread on a test field (1.5l/m2 at 1010 cfu/l) in a drinking water protection zone with loamy to sandy sediments in the Donauried area, Baden-Wurttemberg. Soil samples and groundwaters were collected at monthly intervals October 1994 – June 1995 during which 114 E. coli isolates were recovered. The first occurrence and maximum concentration of E. coli in soil samples taken from more than 20cm depth was in January 1995, declining rapidly with depth and time. All isolates from soil and only one from groundwater showed the RAPD pattern of the spread E. coli strain. The results could not demonstrate a severe negative impact of the spreading of liquid manure on the bacteriological quality of the groundwater in the given geological situation. The distinct strain patterns found in different kinds of liquid manure suggest that genotyping of E. coli by RAPD may be an adequate tool for tracing sources of faecal contamination.

scholarly journals Molecular characterization of clinical carbapenem-resistant Enterobacterales from Qatar

2021 ◽  
Author(s):  
Fatma Ben Abid ◽  
Clement K. M. Tsui ◽  
Yohei Doi ◽  
Anand Deshmukh ◽  
Christi L. McElheny ◽  
...  
Keyword(s):  
Common Species ◽  
Clinical Samples ◽  
Whole Genome ◽  
E Coli ◽  
Carbapenem Resistant ◽  
Genes Encoding ◽  
Encoding Genes ◽  
Sequence Types ◽  
Common Sequence

AbstractOne hundred forty-nine carbapenem-resistant Enterobacterales from clinical samples obtained between April 2014 and November 2017 were subjected to whole genome sequencing and multi-locus sequence typing. Klebsiella pneumoniae (81, 54.4%) and Escherichia coli (38, 25.5%) were the most common species. Genes encoding metallo-β-lactamases were detected in 68 (45.8%) isolates, and OXA-48-like enzymes in 60 (40.3%). blaNDM-1 (45; 30.2%) and blaOXA-48 (29; 19.5%) were the most frequent. KPC-encoding genes were identified in 5 (3.6%) isolates. Most common sequence types were E. coli ST410 (8; 21.1%) and ST38 (7; 18.4%), and K. pneumoniae ST147 (13; 16%) and ST231 (7; 8.6%).

scholarly journals Foreign investment in emerging legal medicinal cannabis markets: the Jamaica case study

2021 ◽  
Vol 17 (1) ◽  
Author(s):  
Marta Rychert ◽  
Machel Anthony Emanuel ◽  
Chris Wilkins
Keyword(s):  
Civil Society ◽  
Foreign Investment ◽  
Production Efficiency ◽  
Production Quality ◽  
Small Scale ◽  
Organic Cultivation ◽  
Enhanced Production ◽  
Political Interference ◽  
Medicinal Cannabis ◽  
The Impact

Abstract Introduction The establishment of a legal market for medicinal cannabis under the Dangerous Drugs Amendment Act 2015 has positioned Jamaica at the forefront of cannabis law reform in the developing world. Many local cannabis businesses have attracted investment from overseas, including from Canada, US and Europe. Aim To explore the opportunities and risks of foreign investment in an emerging domestic legal cannabis market in a developing country. Methods Thematic analysis of semi-structured face-to-face interviews with 22 key informants (KIs) from the Jamaican government, local cannabis industry, academia and civil society, and field observations of legal and illegal cannabis cultivators. Results KIs from the Jamaican public agencies and domestic cannabis entrepreneurs saw foreign investment as an essential source of capital to finance the start-up costs of legal cannabis businesses. Local cannabis entrepreneurs prioritised investors with the greatest financial resources, brand reputation and export networks. They also considered how allied an investor was with their business vision (e.g., organic cultivation, medical vs. recreational). The key benefits of partnering with a foreign investor included transfer of technical knowledge and financial capital, which enhanced production, quality assurance and seed-to-sale tracking. Some KIs expressed concern over investors’ focus on increasing production efficiency and scale at the expense of funding research and development (R&D) and clinical trials. KIs from the local industry, government agencies and civil society highlighted the risks of ‘predatory’ shareholder agreements and domestic political interference. Concerns were raised about the impact of foreign investment on the diversity of the domestic cannabis sector in Jamaica, including the commitment to transition traditional illegal small-scale cannabis cultivators to the legal sector. Conclusion While foreign investment has facilitated the commercialisation of the cannabis sector in Jamaica, regulatory measures are also needed to protect the domestic industry and support the transition of small-scale illegal cultivators to the legal regime. Foreign investments may alter the economic, social and political determinants of health in transitioning from illegal to legal cannabis market economy.

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