Assessment of <i>Escherichia coli</i> B with enhanced permeability to fluorochromes for flow cytometric assays of bacterial cell function

Herrera, Guadalupe; Martínez, Antonio; Blanco, Manuel; O’Connor, José-Enrique

doi:10.1002/cyto.10148

Cited by 35 publications

(33 citation statements)

References 73 publications

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“…Hence, given the photo-transformation of DCFH-DA under sunlight, the addition of DCFH-DA to E. coli should be done only after solar exposure of bacteria in water. Although some researchers used the probe prior to cellular oxidative stress2829, alternatively the probe might be used after if ROS generation is an UV-photo induced process, as reported for cyanobacterium Anabaena sp. under UV-B3031.…”

Section: Resultsmentioning

confidence: 99%

Intracellular mechanisms of solar water disinfection

Castro-Alférez

Polo-López

Fernández‐Ibáñez³

2016

Sci Rep

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Solar water disinfection (SODIS) is a zero-cost intervention measure to disinfect drinking water in areas of poor access to improved water sources, used by more than 6 million people in the world. The bactericidal action of solar radiation in water has been widely proven, nevertheless the causes for this remain still unclear. Scientific literature points out that generation of reactive oxygen species (ROS) inside microorganisms promoted by solar light absorption is the main reason. For the first time, this work reports on the experimental measurement of accumulated intracellular ROS in E. coli during solar irradiation. For this experimental achievement, a modified protocol based on the fluorescent probe dichlorodihydrofluorescein diacetate (DCFH-DA), widely used for oxidative stress in eukaryotic cells, has been tested and validated for E. coli. Our results demonstrate that ROS and their accumulated oxidative damages at intracellular level are key in solar water disinfection.

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Section: Resultsmentioning

confidence: 99%

Intracellular mechanisms of solar water disinfection

Castro-Alférez

Polo-López

Fernández‐Ibáñez³

2016

Sci Rep

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“…B strains have been widely used for mutagenic assays and toxicological studies because they show higher membrane permeability than K-12 strains [20]. Structural studies on the LPS core oligosaccharide have revealed that K-12 strains are devoid of the O antigen while B strains lack the O antigen and the distal part of the core [21].…”

Section: Discussionmentioning

confidence: 99%

Comparative multi-omics systems analysis of Escherichia coli strains B and K-12

et al. 2012

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BackgroundElucidation of a genotype-phenotype relationship is critical to understand an organism at the whole-system level. Here, we demonstrate that comparative analyses of multi-omics data combined with a computational modeling approach provide a framework for elucidating the phenotypic characteristics of organisms whose genomes are sequenced.ResultsWe present a comprehensive analysis of genome-wide measurements incorporating multifaceted holistic data - genome, transcriptome, proteome, and phenome - to determine the differences between Escherichia coli B and K-12 strains. A genome-scale metabolic network of E. coli B was reconstructed and used to identify genetic bases of the phenotypes unique to B compared with K-12 through in silico complementation testing. This systems analysis revealed that E. coli B is well-suited for production of recombinant proteins due to a greater capacity for amino acid biosynthesis, fewer proteases, and lack of flagella. Furthermore, E. coli B has an additional type II secretion system and a different cell wall and outer membrane composition predicted to be more favorable for protein secretion. In contrast, E. coli K-12 showed a higher expression of heat shock genes and was less susceptible to certain stress conditions.ConclusionsThis integrative systems approach provides a high-resolution system-wide view and insights into why two closely related strains of E. coli, B and K-12, manifest distinct phenotypes. Therefore, systematic understanding of cellular physiology and metabolism of the strains is essential not only to determine culture conditions but also to design recombinant hosts.

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“…Recent genome sequence comparisons have revealed that the envelope makeup of E. coli B is quite different from that of K-12 [17,40]. Moreover, it has been demonstrated that the E. coli B strain displays higher membrane permeability than does K-12 [14], as both strains have very different clusters of genes for the synthesis of the core oligosaccharide and O-antigen polysaccharide of LPS. However, a comparative analysis of the envelope proteins in E. coli B and K-12 strains has not been thoroughly studied.…”

Section: Envelope Proteome Reference Maps Of E Coli B Bl21(de3) and mentioning

confidence: 99%

Comparative Analysis of Envelope Proteomes in Escherichia coli B and K-12 Strains

Han

Lee²,

Hong³

2012

J. Microbiol. Biotechnol.

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Recent genome comparisons of E. coli B and K-12 strains have indicated that the makeup of the cell envelopes in these two strains is quite different. Therefore, we analyzed and compared the envelope proteomes of E. coli BL21(DE3) and MG1655. A total of 165 protein spots, including 62 nonredundant proteins, were unambiguously identified by two-dimensional gel electrophoresis and mass spectrometry. Of these, 43 proteins were conserved between the two strains, whereas 4 and 16 strain-specific proteins were identified only in E. coli BL21(DE3) and MG1655, respectively. Additionally, 24 proteins showed more than 2-fold differences in intensities between the B and K-12 strains. The reference envelope proteome maps showed that E. coli envelope mainly contained channel proteins and lipoproteins. Interesting proteomic observations between the two strains were as follows: (i) B produced more OmpF porin with a larger pore size than K-12, indicating an increase in the membrane permeability; (ii) B produced higher amounts of lipoproteins, which facilitates the assembly of outer membrane β-barrel proteins; and (iii) motility-(FliC) and chemotaxis-related proteins (CheA and CheW) were detected only in K-12, which showed that E. coli B is restricted with regard to migration under unfavorable conditions. These differences may influence the permeability and integrity of the cell envelope, showing that E. coli B may be more susceptible than K-12 to certain stress conditions. Thus, these findings suggest that E. coli K-12 and its derivatives will be more favorable strains in certain biotechnological applications, such as cell surface display or membrane engineering studies.

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Assessment of Escherichia coli B with enhanced permeability to fluorochromes for flow cytometric assays of bacterial cell function

Cited by 35 publications

References 73 publications

Intracellular mechanisms of solar water disinfection

Intracellular mechanisms of solar water disinfection

Comparative multi-omics systems analysis of Escherichia coli strains B and K-12

Comparative Analysis of Envelope Proteomes in Escherichia coli B and K-12 Strains

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