scholarly journals Cell Surface Display of Poliovirus Receptor on Escherichia coli, a Novel Method for Concentrating Viral Particles in Water

2011 ◽  
Vol 77 (15) ◽  
pp. 5141-5148 ◽  
Author(s):  
Morteza Abbaszadegan ◽  
Absar Alum ◽  
Hamed Abbaszadegan ◽  
Valerie Stout
Keyword(s):  
Escherichia Coli ◽  
Cell Surface ◽  
Water Samples ◽  
Stop Codon ◽  
Surface Display ◽  
Cell Surface Display ◽  
Bacterial Cells ◽  
Content Type ◽  
E Coli ◽  
Poliovirus Receptor

ABSTRACTThe lack of efficient methods for concentrating viruses in water samples leads to underreporting of viral contamination in source water. A novel strategy for viral concentration was developed using the expression of target virus receptors on bacterial cells. Poliovirus type 1, the most studied enterovirus, was used as a surrogate for enteric viruses. The human poliovirus receptor (hPVR) gene was expressed on the surface ofEscherichia colicells by using the ice nucleation protein (INP) gene. ThehPVRgene was ligated to the 3′ end of theINPgene after the removal of the stop codon. The resulting open reading frame (ORF) was used for the projection of hPVR onto the outer membrane ofE. coli. Gene expression was tested by SDS-PAGE, Western blot, and dot blot analyses, and virion capture ability was confirmed by transmission electron microscopy. The application of engineeredE. colicells for capturing viruses in 1-liter samples of source and drinking water resulted in 75 to 99% procedural recovery efficiency. Cell surface display of viral receptors on bacterial cells opens a new prospect for an efficient and inexpensive alternative tool for capturing and concentrating waterborne viruses in water samples.

scholarly journals Preparation of Sticky Escherichia coli through Surface Display of an Adhesive Catecholamine Moiety

2013 ◽  
Vol 80 (1) ◽  
pp. 43-53 ◽  
Author(s):  
Joseph P. Park ◽  
Min-Jung Choi ◽  
Se Hun Kim ◽  
Seung Hwan Lee ◽  
Haeshin Lee
Keyword(s):  
Escherichia Coli ◽  
Cell Surface ◽  
Communication Systems ◽  
Cell Attachment ◽  
Surface Display ◽  
Content Type ◽  
E Coli ◽  
Material Surfaces ◽  
Mussel Adhesive ◽  
Adhesive Proteins

ABSTRACTMussels attach to virtually all types of inorganic and organic surfaces in aqueous environments, and catecholamines composed of 3,4-dihydroxy-l-phenylalanine (DOPA), lysine, and histidine in mussel adhesive proteins play a key role in the robust adhesion. DOPA is an unusual catecholic amino acid, and its side chain is called catechol. In this study, we displayed the adhesive moiety of DOPA-histidine onEscherichia colisurfaces using outer membrane protein W as an anchoring motif for the first time. Localization of catecholamines on the cell surface was confirmed by Western blot and immunofluorescence microscopy. Furthermore, cell-to-cell cohesion (i.e., cellular aggregation) induced by the displayed catecholamine and synthesis of gold nanoparticles on the cell surface support functional display of adhesive catecholamines. The engineeredE. coliexhibited significant adhesion onto various material surfaces, including silica and glass microparticles, gold, titanium, silicon, poly(ethylene terephthalate), poly(urethane), and poly(dimethylsiloxane). The uniqueness of this approach utilizing the engineered stickyE. coliis that no chemistry for cell attachment are necessary, and the ability of spontaneousE. coliattachment allows one to immobilize the cells on challenging material surfaces such as synthetic polymers. Therefore, we envision that mussel-inspired catecholamine yielded stickyE. colithat can be used as a new type of engineered microbe for various emerging fields, such as whole living cell attachment on versatile material surfaces, cell-to-cell communication systems, and many others.

scholarly journals Functional Cell Surface Display and Controlled Secretion of Diverse Agarolytic Enzymes by Escherichia coli with a Novel Ligation-Independent Cloning Vector Based on the Autotransporter YfaL

2012 ◽  
Vol 78 (9) ◽  
pp. 3051-3058 ◽  
Author(s):  
Hyeok-Jin Ko ◽  
Eunhye Park ◽  
Joseph Song ◽  
Taek Ho Yang ◽  
Hee Jong Lee ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Cell Surface ◽  
Fluorescent Protein ◽  
Surface Display ◽  
Cell Surface Display ◽  
Heterologous Proteins ◽  
Display System ◽  
Reporter Protein ◽  
Content Type ◽  
Ligation Independent Cloning

ABSTRACTAutotransporters have been employed as the anchoring scaffold for cell surface display by replacing their passenger domains with heterologous proteins to be displayed. We adopted an autotransporter (YfaL) ofEscherichia colifor the cell surface display system. The critical regions in YfaL for surface display were identified for the construction of a ligation-independent cloning (LIC)-based display system. The designed system showed no detrimental effect on either the growth of the host cell or overexpressing heterologous proteins on the cell surface. We functionally displayed monomeric red fluorescent protein (mRFP1) as a reporter protein and diverse agarolytic enzymes fromSaccharophagus degradans2-40, including Aga86C and Aga86E, which previously had failed to be functional expressed. The system could display different sizes of proteins ranging from 25.3 to 143 kDa. We also attempted controlled release of the displayed proteins by incorporating a tobacco etch virus protease cleavage site into the C termini of the displayed proteins. The maximum level of the displayed protein was 6.1 × 104molecules per a single cell, which corresponds to 5.6% of the entire cell surface of actively growingE. coli.

scholarly journals Enhanced Display of Lipase on the Escherichia coli Cell Surface, Based on Transcriptome Analysis

2009 ◽  
Vol 76 (3) ◽  
pp. 971-973 ◽  
Author(s):  
Jong Hwan Baek ◽  
Mee-Jung Han ◽  
Seung Hwan Lee ◽  
Sang Yup Lee
Keyword(s):  
Escherichia Coli ◽  
Cell Surface ◽  
Transcriptome Analysis ◽  
Surface Display ◽  
Cell Surface Display ◽  
Display System ◽  
E Coli ◽  
A Cell ◽  
Anchoring Motif ◽  
Cell Surface Display System

ABSTRACT A cell surface display system was developed using Escherichia coli OmpC as an anchoring motif. The fused Pseudomonas fluorescens SIK W1 lipase was successfully displayed on the surface of E. coli cells, and the lipase activity could be enhanced by the coexpression of the gadBC genes identified by transcriptome analysis.

Expressing Xylanases in Escherichia Coli by Cell Surface Display

2013 ◽  
Vol 634-638 ◽  
pp. 965-969
Author(s):  
Mei Na Zhao ◽  
Zongbao Zheng ◽  
Tao Chen
Keyword(s):  
Escherichia Coli ◽  
Cell Surface ◽  
Consolidated Bioprocessing ◽  
Surface Display ◽  
Cell Surface Display ◽  
Whole Cell ◽  
E Coli ◽  
Metabolic Burden ◽  
Whole Cell Biocatalyst ◽  
Anchor Proteins

In this research, xylan was utilized by a recombinant whole cell biocatalyst, which was developed by expressing three xylanases — β-xylosidase, endoxylanase, and α-arabinofuranosidase — on the surface of the E. coli BL21 (DE3). The xylanases were displayed on the surface of the cells by fusing with anchor proteins, Blc. The assimilation of xylan by cell surface display was the first step in the consolidated bioprocessing (CBP). This result shows that the engineering strains could be endowed with the ability to assimilate xylan. The co-display engineering strains utilized xylan and expressed less metabolic burden than the engineering strains secreting extracellular xylanases.

PBP4 and PBP5 are involved in regulating exopolysaccharide synthesis during Escherichia coli biofilm formation

Microbiology ◽  
2021 ◽  
Vol 167 (3) ◽  
Author(s):  
Sathi Mallick ◽  
Shanti Kiran ◽  
Tapas Kumar Maiti ◽  
Anindya S. Ghosh
Keyword(s):  
Escherichia Coli ◽  
Biofilm Formation ◽  
Type Species ◽  
Pentose Phosphate ◽  
Bacterial Cells ◽  
Surface Adhesion ◽  
Content Type ◽  
Penicillin Binding Proteins ◽  
E Coli ◽  
Link Type

Escherichia coli low-molecular-mass (LMM) Penicillin-binding proteins (PBPs) help in hydrolysing the peptidoglycan fragments from their cell wall and recycling them back into the growing peptidoglycan matrix, in addition to their reported involvement in biofilm formation. Biofilms are external slime layers of extra-polymeric substances that sessile bacterial cells secrete to form a habitable niche for themselves. Here, we hypothesize the involvement of Escherichia coli LMM PBPs in regulating the nature of exopolysaccharides (EPS) prevailing in its extra-polymeric substances during biofilm formation. Therefore, this study includes the assessment of physiological characteristics of E. coli CS109 LMM PBP deletion mutants to address biofilm formation abilities, viability and surface adhesion. Finally, EPS from parent CS109 and its ΔPBP4 and ΔPBP5 mutants were purified and analysed for sugars present. Deletions of LMM PBP reduced biofilm formation, bacterial adhesion and their viability in biofilms. Deletions also diminished EPS production by ΔPBP4 and ΔPBP5 mutants, purification of which suggested an increased overall negative charge compared with their parent. Also, EPS analyses from both mutants revealed the appearance of an unusual sugar, xylose, that was absent in CS109. Accordingly, the reason for reduced biofilm formation in LMM PBP mutants may be speculated as the subsequent production of xylitol and a hindrance in the standard flow of the pentose phosphate pathway.

scholarly journals Bacterial Secretions of Nonpathogenic Escherichia coli Elicit Inflammatory Pathways: a Closer Investigation of Interkingdom Signaling

mBio ◽  
2015 ◽  
Vol 6 (2) ◽  
Author(s):  
Amin Zargar ◽  
David N. Quan ◽  
Karen K. Carter ◽  
Min Guo ◽  
Herman O. Sintim ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Epithelial Cells ◽  
Negative Feedback ◽  
Cell Contact ◽  
Bacterial Cells ◽  
Content Type ◽  
Phenotypic Differences ◽  
E Coli ◽  
Autoinducer 2 ◽  
Human Epithelial Cells

ABSTRACTThere have been many studies on the relationship between nonpathogenic bacteria and human epithelial cells; however, the bidirectional effects of the secretomes (secreted substances in which there is no direct bacterium-cell contact) have yet to be fully investigated. In this study, we use a transwell model to explore the transcriptomic effects of bacterial secretions from two different nonpathogenicEscherichia colistrains on the human colonic cell line HCT-8 using next-generation transcriptome sequencing (RNA-Seq).E. coliBL21 and W3110, while genetically very similar (99.1% homology), exhibit key phenotypic differences, including differences in their production of macromolecular structures (e.g., flagella and lipopolysaccharide) and in their secretion of metabolic byproducts (e.g., acetate) and signaling molecules (e.g., quorum-sensing autoinducer 2 [AI-2]). After analysis of differential epithelial responses to the respective secretomes, this study shows for the first time that a nonpathogenic bacterial secretome activates the NF-κB-mediated cytokine-cytokine receptor pathways while also upregulating negative-feedback components, including the NOD-like signaling pathway. Because of AI-2's relevance as a bacterium-bacterium signaling molecule and the differences in its secretion rates between these strains, we investigated its role in HCT-8 cells. We found that the expression of the inflammatory cytokine interleukin 8 (IL-8) responded to AI-2 with a pattern of rapid upregulation before subsequent downregulation after 24 h. Collectively, these data demonstrate that secreted products from nonpathogenic bacteria stimulate the transcription of immune-related biological pathways, followed by the upregulation of negative-feedback elements that may serve to temper the inflammatory response.IMPORTANCEThe symbiotic relationship between the microbiome and the host is important in the maintenance of human health. There is a growing need to further understand the nature of these relationships to aid in the development of homeostatic probiotics and also in the design of novel antimicrobial therapeutics. To our knowledge, this is the first global-transcriptome study of bacteria cocultured with human epithelial cells in a model to determine the transcriptional effects of epithelial cells in which epithelial and bacterial cells are allowed to “communicate” with each other only through diffusible small molecules and proteins. By beginning to demarcate the direct and indirect effects of bacteria on the gastrointestinal (GI) tract, two-way interkingdom communication can potentially be mediated between host and microbe.

scholarly journals Bioinformatic and Molecular Analysis of Inverse Autotransporters from Escherichia coli

mSphere ◽  
2019 ◽  
Vol 4 (4) ◽  
Author(s):  
Kelvin G. K. Goh ◽  
Danilo G. Moriel ◽  
Steven J. Hancock ◽  
Minh-Duy Phan ◽  
Mark A. Schembri
Keyword(s):  
Escherichia Coli ◽  
Cell Surface ◽  
Biofilm Formation ◽  
Secretion System ◽  
Content Type ◽  
E Coli ◽  
Wide Range ◽  
Type V Secretion ◽  
K 12 ◽  
Type V

ABSTRACT Proteins secreted by the type V secretion system possess multiple functions, including the capacity to mediate adhesion, aggregation, and biolfilm formation. The type V secretion system can be divided into five subclasses, one of which is the type Ve system. Proteins of the type Ve secretion system are also referred to as inverse autotransporters (IATs). In this study, we performed an in silico analysis of 126 completely sequenced Escherichia coli genomes available in the NCBI database and identified several distinct IAT-encoding gene families whose distribution varied throughout the E. coli phylogeny. The genes included three characterized IATs (intimin, fdeC, and yeeJ) and four uncharacterized IATs (here named iatA, iatB, iatC, and iatD). The four iat genes were cloned from the completely sequenced environmental E. coli strain SMS-3-5 and characterized. Three of these IAT proteins (IatB, IatC, and IatD) were expressed at the cell surface and possessed the capacity to mediate biofilm formation in a recombinant E. coli K-12 strain. Further analysis of the iatB gene, which showed a unique association with extraintestinal E. coli strains, suggested that its regulation is controlled by the LeuO global regulator. Overall, this study provides new data describing the prevalence, sequence variation, domain structure, function, and regulation of IATs found in E. coli. IMPORTANCE Escherichia coli is one of the most prevalent facultative anaerobes of the human gut. E. coli normally exists as a harmless commensal but can also cause disease following the acquisition of genes that enhance its pathogenicity. Adhesion is an important first step in colonization of the host and is mediated by an array of cell surface components. In E. coli, these include a family of adhesins secreted by the type V secretion system. Here, we identified and characterized new proteins from an emerging subclass of the type V secretion system known as the inverse autotransporters (IATs). We found that IAT-encoding genes are present in a wide range of strains and showed that three novel IATs were localized on the E. coli cell surface and mediated biofilm formation. Overall, this study provides new insight into the prevalence, function, and regulation of IATs in E. coli.

Changes in Cell Surface Properties of Shiga Toxigenic Escherichia coli by Quercus infectoria G. Olivier

2009 ◽  
Vol 72 (8) ◽  
pp. 1699-1704 ◽  
Author(s):  
SUPAYANG PIYAWAN VORAVUTHIKUNCHAI ◽  
SAKOL SUWALAK
Keyword(s):  
Escherichia Coli ◽  
Cell Surface ◽  
Surface Properties ◽  
Cell Surface Hydrophobicity ◽  
Ethanolic Extract ◽  
Bacterial Cells ◽  
Electron Microscopic ◽  
Cell Surface Properties ◽  
E Coli ◽  
Quercus Infectoria

The effects of Quercus infectoria (family Fagaceae) nutgalls on cell surface properties of Shiga toxigenic Escherichia coli (STEC) were investigated with an assay of microbial adhesion to hydrocarbon. The surface of bacterial cells treated with Q. infectoria exhibited a higher level of cell surface hydrophobicity (CSH) toward toluene than did the surface of untreated cells. With 50% ethanolic extract, the CSH of the three strains of STEC O157:H7 treated with 4× MIC of the extract resulted in moderate or strong hydrophobicity, whereas at 2× MIC and MIC, the CSH of only one strain of E. coli O157:H7 was significantly affected. The 95% ethanolic extract had a significant effect on CSH of all three strains at both 4× MIC and 2× MIC but not at the MIC. The effect on bacterial CSH was less pronounced with the other STEC strains. At 4× MIC, the 50% ethanolic extract increased the CSH of all non-O157 STEC strains significantly. At 2× MIC and 4× MIC, the 95% ethanolic extract affected the CSH of E. coli O26:H11 significantly but did not affect E. coli O111:NM or E. coli O22. Electron microscopic examination revealed the loss of pili in the treated cells. The ability of Q. infectoria extract to modify hydrophobic domains enables this extract to partition the lipids of the bacterial cell membrane, rendering the membrane more permeable and allowing leakage of ions and other cell contents, which leads to cell death. Further studies are required to evaluate the effects of Q. infectoria extract in food systems or in vivo and provide support for the use of this extract as a food additive for control of these STEC pathogens.

scholarly journals Impacts of Hematite Nanoparticle Exposure on Biomechanical, Adhesive, and Surface Electrical Properties of Escherichia coli Cells

2012 ◽  
Vol 78 (11) ◽  
pp. 3905-3915 ◽  
Author(s):  
Wen Zhang ◽  
Joseph Hughes ◽  
Yongsheng Chen
Keyword(s):  
Escherichia Coli ◽  
Electrical Properties ◽  
Current Knowledge ◽  
Statistical Significance ◽  
Spring Constant ◽  
Bacterial Cells ◽  
Kelvin Probe ◽  
Content Type ◽  
Force Microscopy ◽  
E Coli

ABSTRACTDespite a wealth of studies examining the toxicity of engineered nanomaterials, current knowledge on their cytotoxic mechanisms (particularly from a physical perspective) remains limited. In this work, we imaged and quantitatively characterized the biomechanical (hardness and elasticity), adhesive, and surface electrical properties ofEscherichia colicells with and without exposure to hematite nanoparticles (NPs) in an effort to advance our understanding of the cytotoxic impacts of nanomaterials. Both scanning electron microscopy (SEM) and atomic force microscopy (AFM) showed thatE. colicells had noticeable deformation with hematite treatment for 45 min with a statistical significance. The hematite-treated cells became significantly harder or stiffer than untreated ones, as evidenced by indentation and spring constant measurements. The average indentation of the hematite-treatedE. colicells was 120 nm, which is significantly lower (P< 0.01) than that of the untreated cells (approximately 400 nm). The spring constant of hematite-treatedE. colicells (0.28 ± 0.11 nN/nm) was about 20 times higher than that of untreated ones (0.01 ± 0.01 nN/nm). The zeta potential ofE. colicells, measured by dynamic light scattering (DLS), was shown to shift from −4 ± 2 mV to −27 ± 8 mV with progressive surface adsorption of hematite NPs, a finding which is consistent with the local surface potential measured by Kelvin probe force microscopy (KPFM). Overall, the reported findings quantitatively revealed the adverse impacts of nanomaterial exposure on physical properties of bacterial cells and should provide insight into the toxicity mechanisms of nanomaterials.

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