scholarly journals Structural and Topographical Studies of the Type IV Bundle-Forming Pilus Assembly Complex of Enteropathogenic Escherichia coli

2003 ◽  
Vol 185 (22) ◽  
pp. 6695-6701 ◽  
Author(s):  
Jaiweon Hwang ◽  
David Bieber ◽  
Sandra W. Ramer ◽  
Cheng-Yen Wu ◽  
Gary K. Schoolnik
Keyword(s):  
Escherichia Coli ◽  
Specific Antibodies ◽  
Type Iv ◽  
Transmission Electron ◽  
Human Volunteers ◽  
Oligomeric Complex ◽  
And Function

ABSTRACT The type IV bundle-forming pili (BFP) of enteropathogenic Escherichia coli (EPEC) are required for virulence in orally challenged human volunteers and for the localized adherence and autoaggregation in vitro phenotypes. BFP filament biogenesis and function are encoded by the 14-gene bfp operon. The BFP assembly complex, containing a BfpB-His6 fusion protein, was chemically cross-linked in situ, and the complex was then purified from BFP-expressing EPEC by a combination of nickel- and BfpB antibody-based affinity chromatography. Characterization of the isolated complex by immunoblotting using BFP protein-specific antibodies showed that at least 10 of the 14 proteins specified by the bfp operon physically interact to form an oligomeric complex. Proteins localized to the outer membrane, inner membrane, and periplasm are within this complex, thus demonstrating that the complex spans the periplasmic space. A combination of immunofluorescence and immuno-gold thin-section transmission electron microscopy studies localized this complex to one pole of the cell.

Drosophila PS integrins recognize vertebrate vitronectin and function as cell-substratum adhesion receptors in vitro

Development ◽  
1991 ◽  
Vol 113 (3) ◽  
pp. 1007-1016 ◽  
Author(s):  
S. Hirano ◽  
K. Ui ◽  
T. Miyake ◽  
T. Uemura ◽  
M. Takeichi
Keyword(s):  
Cell Attachment ◽  
Collagen Type I ◽  
Matrix Proteins ◽  
Type I ◽  
Rgd Peptides ◽  
Drosophila Cell ◽  
Type Iv ◽  
And Function

Using the Drosophila cell line MLDmBG-1, a monoclonal antibody aBG-1 that can inhibit not only cell clumping but also cell spreading was generated. This antibody immunoprecipitates a complex of molecules consisting of a major 120 × 10(3) Mr and other components. To characterize the 120 × 10(3) Mr component, we purified it, generated antibodies to it, and cloned its cDNA. Sequencing of this cDNA suggests that the 120 × 10(3) Mr molecule is identical to PS beta, a beta chain of Drosophila integrins. The other components immunoprecipitated included two alpha chains of Drosophila integrins, PS1 alpha and PS2 alpha, as revealed using specific antibodies to these molecules. These suggest that aBG-1 recognizes the PS beta associated with PS1 alpha or PS2 alpha. However, immunostaining of embryos and larvae with aBG-1 showed that the staining pattern is similar to that for PS2 alpha but not for PS beta, suggesting that the antibody preferentially recognizes the PS beta associated with particular alpha chains in situ. We then attempted to characterize the ligands for these integrin complexes, using culture dishes coated with various vertebrate matrix proteins. These cells spread very well on dishes coated with vitronectin and, to a lesser extent, on those with fibronectin. This spreading was partially inhibited by aBG-1, but not by other control antibodies or RGD peptides. The cell attachment to these substrata was not affected by the antibody. The cells also can attach to dishes coated with laminin but without spreading, and this attachment was not inhibited by aBG-1. Furthermore, they do not attach to dishes coated with collagen type I, type IV, and fibrinogen. These results indicate that Drosophila PS integrins can recognize vertebrate vitronectin, and also fibronectin with a weaker affinity, at sites other than RGD sequences, and thus can function in cell-substratum adhesion.

Preparation and Characterization of Fibrous Hydroxyapatite/Chitosan Nanocomposites with High Hydroxyapatite Dosage

2012 ◽  
Vol 457-458 ◽  
pp. 365-371 ◽  
Author(s):  
Cai Yun Zhang ◽  
Dai Yin Peng ◽  
Chuan Hua Lu ◽  
Xian Ping Wang ◽  
Qian Feng Fang
Keyword(s):  
Morphological Properties ◽  
In Vitro Tests ◽  
X Ray Diffraction ◽  
Permeable Membrane ◽  
X Ray ◽  
Chitosan Matrix ◽  
Transmission Electron

In this paper the hydroxyapatite fibers reinforced chitosan nanocomposites with high hydroxyapatite dosage (70~90 wt%) were synthesized by in-situ hybridization. The semi-permeable membrane was used to control the process of hybridization and morphology of hydroxyapatite. The compositional and morphological properties of nanocomposites were investigated by FTIR spectroscopy, X-ray diffraction, and transmission electron microscopy. The results showed that the hydroxyapatite were carbonated nanometer crystalline fibers with high aspect ratio (about 25) and dispersed uniformly in the nanocomposites. The high-resolution image indicated that the growth of nano-hydroxyapatite crystallites in the chitosan matrix preferred in the c-axis. The mechanical properties of these nanocomposites were enhanced dramatically and the compressive strength increases almost to 170MPa when the hydroxyapatite content is 70 wt%. The in vitro tests indicated that the composites have high bioactivity and degradation. These properties illustrated the potential application of this kind of nanocomposites for bone tissue engineering.

scholarly journals Characterization of a Novel Type IV Pilus Locus Encoded on the Large Plasmid of Locus of Enterocyte Effacement-Negative Shiga-Toxigenic Escherichia coli Strains That Are Virulent for Humans

2002 ◽  
Vol 70 (6) ◽  
pp. 3094-3100 ◽  
Author(s):  
Potjanee Srimanote ◽  
Adrienne W. Paton ◽  
James C. Paton
Keyword(s):  
Escherichia Coli ◽  
Gastrointestinal Disease ◽  
Large Plasmid ◽  
Type Iv Pilus ◽  
Upstream Gene ◽  
Type Iv ◽  
Uremic Syndrome ◽  
Enterocyte Effacement

ABSTRACT The majority of Shiga-toxigenic Escherichia coli (STEC) strains isolated from humans with gastrointestinal disease carry large (approximately 90-kb) plasmids. We have been analyzing the megaplasmid (designated pO113) from an O113:H21 STEC strain (98NK2). This strain lacks the locus for enterocyte effacement (LEE) and yet was responsible for an outbreak of hemolytic uremic syndrome. In the present study, we demonstrate that pO113 carries a novel type IV pilus biosynthesis locus (pil) related to those of the IncI plasmids R721, R64, and ColIb9. The pO113 pil locus consists of 11 closely linked genes (pilL through pilV) with an additional separately transcribed upstream gene (pilI). It directs the expression of long thin pili on the 98NK2 surface and the hemagglutination of guinea pig erythrocytes. We also demonstrate that pO113 can be transferred by conjugation. However, the type IV pilus encoded by pO113 does not appear to be involved in the adherence of 98NK2 to HEp-2 or Hct-8 cells in vitro. Homologues of the pO113 pil locus were present in several other LEE-negative STEC strains but not in LEE-positive STEC strains belonging to serogroup O26, O111, or O157.

The Use of Advanced Analytical Techniques for Characterization of the Chemical, Physical, and Crystallographic Nature of a Surface

1973 ◽  
Vol 31 ◽  
pp. 132-133 ◽  
Author(s):  
R. E. Herfert
Keyword(s):  
Surface Characterization ◽  
Analytical Techniques ◽  
X Ray Diffraction ◽  
Depth Of Penetration ◽  
X Ray ◽  
The Past ◽  
Transmission Electron ◽  
Scanning Electron

Studies of the nature of a surface, either metallic or nonmetallic, in the past, have been limited to the instrumentation available for these measurements. In the past, optical microscopy, replica transmission electron microscopy, electron or X-ray diffraction and optical or X-ray spectroscopy have provided the means of surface characterization. Actually, some of these techniques are not purely surface; the depth of penetration may be a few thousands of an inch. Within the last five years, instrumentation has been made available which now makes it practical for use to study the outer few 100A of layers and characterize it completely from a chemical, physical, and crystallographic standpoint. The scanning electron microscope (SEM) provides a means of viewing the surface of a material in situ to magnifications as high as 250,000X.

scholarly journals Enlisting the Ixodes scapularis Embryonic ISE6 Cell Line to Investigate the Neuronal Basis of Tick—Pathogen Interactions

Pathogens ◽  
2021 ◽  
Vol 10 (1) ◽  
pp. 70
Author(s):  
Lourdes Mateos-Hernández ◽  
Natália Pipová ◽  
Eléonore Allain ◽  
Céline Henry ◽  
Clotilde Rouxel ◽  
...  
Keyword(s):  
Cell Line ◽  
Peripheral Nerves ◽  
Ixodes Scapularis ◽  
Embryonic Cell ◽  
Cell Subpopulation ◽  
In Vivo Experiments

Neuropeptides are small signaling molecules expressed in the tick central nervous system, i.e., the synganglion. The neuronal-like Ixodes scapularis embryonic cell line, ISE6, is an effective tool frequently used for examining tick–pathogen interactions. We detected 37 neuropeptide transcripts in the I. scapularis ISE6 cell line using in silico methods, and six of these neuropeptide genes were used for experimental validation. Among these six neuropeptide genes, the tachykinin-related peptide (TRP) of ISE6 cells varied in transcript expression depending on the infection strain of the tick-borne pathogen, Anaplasma phagocytophilum. The immunocytochemistry of TRP revealed cytoplasmic expression in a prominent ISE6 cell subpopulation. The presence of TRP was also confirmed in A. phagocytophilum-infected ISE6 cells. The in situ hybridization and immunohistochemistry of TRP of I. scapularis synganglion revealed expression in distinct neuronal cells. In addition, TRP immunoreaction was detected in axons exiting the synganglion via peripheral nerves as well as in hemal nerve-associated lateral segmental organs. The characterization of a complete Ixodes neuropeptidome in ISE6 cells may serve as an effective in vitro tool to study how tick-borne pathogens interact with synganglion components that are vital to tick physiology. Therefore, our current study is a potential stepping stone for in vivo experiments to further examine the neuronal basis of tick–pathogen interactions.

scholarly journals Characterization of a Dye-Decolorizing Peroxidase from Irpex lacteus Expressed in Escherichia coli: An Enzyme with Wide Substrate Specificity Able to Transform Lignosulfonates

2021 ◽  
Vol 7 (5) ◽  
pp. 325
Author(s):  
Laura Isabel de de Eugenio ◽  
Rosa Peces-Pérez ◽  
Dolores Linde ◽  
Alicia Prieto ◽  
Jorge Barriuso ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Inclusion Bodies ◽  
Veratryl Alcohol ◽  
Dye Decolorization ◽  
Irpex Lacteus ◽  
Type D ◽  
Lignin Peroxidases

A dye-decolorizing peroxidase (DyP) from Irpex lacteus was cloned and heterologously expressed as inclusion bodies in Escherichia coli. The protein was purified in one chromatographic step after its in vitro activation. It was active on ABTS, 2,6-dimethoxyphenol (DMP), and anthraquinoid and azo dyes as reported for other fungal DyPs, but it was also able to oxidize Mn2+ (as manganese peroxidases and versatile peroxidases) and veratryl alcohol (VA) (as lignin peroxidases and versatile peroxidases). This corroborated that I. lacteus DyPs are the only enzymes able to oxidize high redox potential dyes, VA and Mn+2. Phylogenetic analysis grouped this enzyme with other type D-DyPs from basidiomycetes. In addition to its interest for dye decolorization, the results of the transformation of softwood and hardwood lignosulfonates suggest a putative biological role of this enzyme in the degradation of phenolic lignin.

scholarly journals Cryo-ET Characterization of Novel Cellular Extrusions in Escherichia coli Induced by the Major Subunit Protein of Type IV Pili, PilA, from Pseudomonas aeruginosa

2021 ◽  
Vol 27 (S1) ◽  
pp. 280-282
Author(s):  
Juan Sanchez ◽  
Daniel Parrell ◽  
Alba Gonzalez-Rivera ◽  
Nicoleta Ploscariu ◽  
Katrina Forest ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Pseudomonas Aeruginosa ◽  
Type Iv Pili ◽  
Subunit Protein ◽  
Type Iv

scholarly journals Isolation and characterization of pathogenic Escherichia coli bacteriophages from chicken and beef offal

2020 ◽  
Vol 13 (1) ◽  
Author(s):  
Celosia Lukman ◽  
Christopher Yonathan ◽  
Stella Magdalena ◽  
Diana Elizabeth Waturangi
Keyword(s):  
Escherichia Coli ◽  
Foodborne Pathogens ◽  
High Efficiency ◽  
Multiplicity Of Infection ◽  
Isolation And Characterization ◽  
Transmission Electron ◽  
Pathogenic Escherichia Coli ◽  
Family Based

Abstract Objective This study was conducted to isolate and characterize lytic bacteriophages for pathogenic Escherichia coli from chicken and beef offal, and analyze their capability as biocontrol for several foodborne pathogens. Methods done in this research are bacteriophage isolation, purification, titer determination, application, determination of host range and minimum multiplicity of infection (miMOI), and bacteriophage morphology. Results Six bacteriophages successfully isolated from chicken and beef offal using EPEC and EHEC as host strain. Bacteriophage titers observed between 109 and 1010 PFU mL−1. CS EPEC and BL EHEC bacteriophage showed high efficiency in reduction of EPEC or EHEC contamination in meat about 99.20% and 99.04%. The lowest miMOI was 0.01 showed by CS EPEC bacteriophage. CI EPEC and BL EPEC bacteriophage suspected as Myoviridae family based on its micrograph from Transmission Electron Microscopy (TEM). Refers to their activity, bacteriophages isolated in this study have a great potential to be used as biocontrol against several foodborne pathogens.

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