Pseudomonas aeruginosa is an opportunistic gram-negative bacterium implicated in acute and chronic nosocomial infections and a leading cause of patient mortality. Such infections occur owing to biofilm formation that confers multidrug resistance and enhanced pathogenesis to the bacterium. In this study, we used a rational drug design strategy to inhibit the quorum signaling system of P. aeruginosa by designing potent inhibitory lead molecules against anthranilate-CoA ligase enzyme encoded by the pqsA gene. This enzyme produces autoinducers for cell-to-cell communication, which result in biofilm formation, and thus plays a pivotal role in the virulence of P. aeruginosa. A library of potential drug molecules was prepared by performing ligand-based screening using an available set of enzyme inhibitors. Subsequently, structure-based virtual screening was performed to identify compounds showing the best binding conformation with the target enzyme and forming a stable complex. The two hit compounds interact with the binding site of the enzyme through multiple short-range hydrophilic and hydrophobic interactions. Molecular dynamic simulation and MM-PBSA/GBSA results to calculate the affinity and stability of the hit compounds with the PqsA enzyme further confirmed their strong interactions. The hit compounds might be useful in tackling the resistant phenotypes of this pathogen.

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Toxic heavy metal accumulation is one of anthropogenic environmental pollutions, which poses risks to human health and ecological systems. Conventional heavy metal remediation approaches rely on expensive chemical and physical processes leading to the formation and release of other toxic waste products. Instead, microbial bioremediation has gained interest as a promising and cost-effective alternative to conventional methods, but the genetic complexity of microorganisms and the lack of appropriate genetic engineering technologies have impeded the development of bioremediating microorganisms. Recently, the emerging synthetic biology opened a new avenue for microbial bioremediation research and development by addressing the challenges and providing novel tools for constructing bacteria with enhanced capabilities: rapid detection and degradation of heavy metals while enhanced tolerance to toxic heavy metals. Moreover, synthetic biology also offers new technologies to meet biosafety regulations since genetically modified microorganisms may disrupt natural ecosystems. In this review, we introduce the use of microorganisms developed based on synthetic biology technologies for the detection and detoxification of heavy metals. Additionally, this review explores the technical strategies developed to overcome the biosafety requirements associated with the use of genetically modified microorganisms.

Background Industrial biofuels and other value-added products can be produced from metabolically engineered microorganisms. Methylomonas sp. DH-1 is a candidate platform for bioconversion that uses methane as a carbon source. Although several genetic engineering techniques have been developed to work with Methylomonas sp. DH-1, the genetic manipulation of plasmids remains difficult because of the restriction-modification (RM) system present in the bacteria. Therefore, the RM system in Methylomonas sp. DH-1 must be identified to improve the genetic engineering prospects of this microorganism. Results We identified a DNA methylation site, TGGCCA, and its corresponding cytosine methyltransferase for the first time in Methylomonas sp. DH-1 through whole-genome bisulfite sequencing. The methyltransferase was confirmed to methylate the fourth nucleotide of TGGCCA. In general, methylated plasmids exhibited better transformation efficiency under the protection of the RM system than non-methylated plasmids did. As expected, when we transformed Methylomonas sp. DH-1 with plasmid DNA harboring the psy gene, the metabolic flux towards carotenoid increased. The methyltransferase-treated plasmid exhibited an increase in transformation efficiency of 2.5 × 103 CFU/μg (124%). The introduced gene increased the production of carotenoid by 26%. In addition, the methyltransferase-treated plasmid harboring anti-psy sRNA gene exhibited an increase in transformation efficiency by 70% as well. The production of carotenoid was decreased by 40% when the psy gene was translationally repressed by anti-psy sRNA. Conclusions Plasmid DNA methylated by the discovered cytosine methyltransferase from Methylomonas sp. DH-1 had a higher transformation efficiency than non-treated plasmid DNA. The RM system identified in this study may facilitate the plasmid-based genetic manipulation of methanotrophs.

Lycopene is a carotenoid widely used as a food and feed supplement due to its antioxidant, anti-inflammatory, and anti-cancer functions. Various metabolic engineering strategies have been implemented for high lycopene production in Escherichia coli , and for this purpose it was essential to select and develop an E. coli strain with the highest potency. In this study, we evaluated 16 E. coli strains to determine the best lycopene production host by introducing a lycopene biosynthetic pathway ( crtE , crtB , and crtI genes cloned from Deinococcus wulumuqiensis R12 and dxs , dxr , ispA , and idi genes cloned from E. coli ). The 16 lycopene strain titers diverged from 0 to 0.141 g/l, with MG1655 demonstrating the highest titer (0.141 g/l), while the SURE and W strains expressed the lowest (0 g/l) in an LB medium. When a 2 × YTg medium replaced the MG1655 culture medium, the titer further escalated to 1.595 g/l. These results substantiate that strain selection is vital in metabolic engineering, and further, that MG1655 is a potent host for producing lycopene and other carotenoids with the same lycopene biosynthetic pathway.