A modified Pseudomonas aeruginosa strain capable of overexpressing the estA gene, an encoding gene for a membrane-bound esterase, was constructed and its rhamnolipid (RML) production was studied. Fermentations using wild-type (WT) and modified P. aeruginosa strains were conducted until exhaustion of glycerol in Medium Salt Production, using two different C/N ratios. At a C/N of 83.2, the modified strain produced up to 3.9 times more RMLs than the WT, yielding a maximum concentration of 14.62 g/L RML when measured by HPLC and 22 g/L by the orcinol assay. Cell-free supernatant from the modified strain reduced surface tension to 29.4 mN/m and had a CMC of 240 mg/L and CMD of 56.05. This is the first report on the construction of an estA-based recombinant strain for RML production.
The need to find more stable catalysts has encouraged the study of naturally resilient enzymes, such as those found in extremophile organisms. In the present work, the influence of rare codons on the expression in Escherichia coli of the lipase Pf2001Δ60 from Pyrococcus furiosus was evaluated. Expression was carried out in two E. coli strains, BL21(DE3)pLysS and the rare tRNA supplemented Rosetta(DE3)pLysS. 3(2) factorial design was used to appraise the influence of temperature and inducer concentration on enzyme expression every hour for the 4-h expression period. Four response surfaces were constructed for each time, and the statistical parameters were evaluated. Lipase production was twice as high for Rosetta(DE3)pLysS than for BL21(DE3)pLysS. The factorial design indicated that optimal expression occurred at 30 °C after 4h, with lipase production of 240 U/L. The analysis of statistical parameters during the expression time showed that the velocity at which the enzyme was produced affected cell growth and maximum activity, with a higher speed leading to lower expression and cell growth. The presence of rare tRNAs prevented bottlenecks in lipase expression, and the experimental design was shown to be important for maximizing the production strategies and minimizing the metabolic load to which the host is subjected.
Surfactin, a biosurfactant with great activity on interfaces, have been reported as a great substitute to non-renewable sources, non-biologically synthesized surfactants. It is expected to see more studies at the next years evolving its application, including on marine environments, especially ones impacted with petroleum or other contaminants. In this review we address in details the main aspects of surfactin production, including main microorganisms, cultivation modes, pathways and conditions. We address the main aspects of surfactin production by Bacillus subtilis with the different strategies explored to reach this bioprocess up to large scale, as well as the main challenges encountered. As well, is detailed its recovery and purification methods, that generally combine two or more steps as acid precipitation, solvent extraction, liquid membrane extraction, foam fractionation and membrane-based techniques. We also provide a brief summary of its potential application on marine environments, and our prospects from future application, as a brief outlook on physiochemistry characteristics of the main molecules.
Surfactin, a biosurfactant with great activity on interfaces, have been reported as a great substitute to non-renewable sources, non-biologically synthesized surfactants. It is expected to see more studies at the next years evolving its application, including on marine environments, especially ones impacted with petroleum or other contaminants. In this review we address in details the main aspects of surfactin production, including main microorganisms, cultivation modes, pathways and conditions. We address the main aspects of surfactin production by Bacillus subtilis with the different strategies explored to reach this bioprocess up to large scale, as well as the main challenges encountered. As well, is detailed its recovery and purification methods, that generally combine two or more steps as acid precipitation, solvent extraction, liquid membrane extraction, foam fractionation and membrane-based techniques. We also provide a brief summary of its potential application on marine environments, and our prospects from future application, as a brief outlook on physiochemistry characteristics of the main molecules.
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