scholarly journals Generation of transducible version of a recombinant human HAND2 transcription factor from Escherichia coli

2021 ◽  
Author(s):  
Krishna Kumar Haridhasapavalan ◽  
Pradeep Kumar Sundaravadivelu ◽  
Anshuman Mohapatra ◽  
Neha Joshi ◽  
Nayan Jyoti Das ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Transcription Factor ◽  
Nuclear Translocation ◽  
Affinity Purification ◽  
Cell Penetration ◽  
Protein Coding ◽  
E Coli ◽  
Heterologous System ◽  
One Step ◽  
The One

AbstractTranscription factor HAND2 has a significant role in vascularization, angiogenesis, and cardiac neural crest development. Also, it is one of the key cardiac factors crucial for the enhanced derivation of functional and mature myocytes from non-myocyte cells. Here, we report the generation of the recombinant human HAND2 fusion protein from the heterologous system. First, we cloned the full-length human HAND2 gene (only protein-coding sequence) after codon optimization along with the fusion tags (for cell penetration, nuclear translocation, and affinity purification) into the expression vector. We then transformed and expressed it in Escherichia coli (E. coli) strain, BL21(DE3). Next, the effect (in terms of expression) of tagging of fusion tags with this recombinant protein at two different terminals was also investigated. Notably, using affinity chromatography, we established the one-step homogeneous purification of human recombinant HAND2 protein; and through circular dichroism spectroscopy, we established that this purified protein had retained its secondary structure. Furthermore, we show that this purified human protein could transduce the human cells and translocate to its nucleus. Prospectively, the purified recombinant HAND2 protein can potentially be a safe and effective molecular tool in the direct cardiac reprogramming process and other biological applications.

scholarly journals Effect of a lytic bacteriophage on rabbits experimentally infected with pathogenic Escherichia coli

2017 ◽  
Vol 25 (3) ◽  
pp. 273 ◽  
Author(s):  
J. Zhao ◽  
L. He ◽  
L. Pan ◽  
Y. Liu ◽  
H. Yao ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Burst Size ◽  
Lethal Dose ◽  
Resistant Bacteria ◽  
Lytic Bacteriophage ◽  
Antibiotic Resistant ◽  
E Coli ◽  
Pathogenic Escherichia Coli ◽  
One Step ◽  
The One

Pathogenic <em>Escherichia coli</em> (<em>E. coli</em>) is severely threatening the rabbit industry in China, and the concern over antibiotic-resistant bacteria has given rise to an urgent need for antibiotic alternatives. In this study, a member (ZRP1) of the <em>Myoviridae</em> family was isolated from rabbit faeces using a strain of rabbit atypical enteropathogenic <em>E. coli</em> (ZR1) as host. The one-step growth curve indicated that the latent period was around 25 to 30 min and the burst size was 144±31 plaque-forming unit/cell. The rate of phage-resistant mutation was 7×10<sup>–5</sup>±4×10<sup>–5</sup>. When the bacteriophage input at the multiplicity of infection (MOI) was 0.1, 1 or 10, the growth of host <em>E. coli</em> in broth was inhibited for 5 h. A single intravenous injection of ZRP1 at MOI 0.1, 1 or 10 significantly prolonged the survival time of rabbits which simultaneously received a lethal dose of ZR1.

scholarly journals Isolation and Characterization of ΦGF1, a Morphotype C3 Bacteriophage that InfectsEscherichia coli

2019 ◽  
Author(s):  
Renzo Punil ◽  
Miguel Talledo ◽  
Mayra Arcondo ◽  
Katherine Suárez ◽  
Kattya Zumaeta
Keyword(s):  
Escherichia Coli ◽  
Burst Size ◽  
Treatment Plant ◽  
Lytic Bacteriophage ◽  
Bacterial Populations ◽  
E Coli ◽  
Isolation And Characterization ◽  
Short Latent Period ◽  
One Step ◽  
The One

ABSTRACTIt has been isolated a lytic bacteriophage specific toEscherichia coli, which can infect at least one different bacterial group. Phage ФGF1 was isolated from a wastewater treatment plant. It is resistant to the effect of chloroform and is stable at 40 and 50 °C. In addition, it is stable in the range of pH 5-8. Its host range is wide, infecting even strains from another genus such asShigella. The one-step growth curve yielded a short latent period of 15 minutes and a burst size of 85 PFU per infected cell. Under the electron microscope, this phage presents the C3 morphotype, extremely rare among members of the Podoviridae family. Phage ФGF1 shows some characteristics that could be considered useful in biocontrol applications againstE. coli. Keywords: Bacteriophage,Escherichia coli, morphotype C3,Podoviridae.IMPORTANCEWastewater throughout the world is a heavy carrier of potential pathogens that live in their environment along with other biological agents, such as bacteriophages, which play a controlling role of the bacterial populations there, as in soil. The description of the diversity of such bacteriophages is of paramount importance since they could be used to intentionally reduce or remove those pathogens from that environment. Our work describes a bacteriophage that lives primarily in this type of water.

One-Step Preparation of Competent E. coli: Transformation and Storage of Bacterial Cells in the Same Solution

2021 ◽  
Vol 2021 (11) ◽  
pp. pdb.prot101212 ◽  
Author(s):  
Michael R. Green ◽  
Joseph Sambrook
Keyword(s):  
Escherichia Coli ◽  
Convenient Method ◽  
Plasmid Dna ◽  
Transformation Efficiency ◽  
Bacterial Cells ◽  
E Coli ◽  
One Step ◽  
And Storage

This protocol describes a convenient method for the preparation, use, and storage of competent Escherichia coli. The reported transformation efficiency of this method is ∼5 × 107 transformants/µg of plasmid DNA.

scholarly journals mcr-1 Gene Expression Modulates the Inflammatory Response of Human Macrophages to Escherichia coli

2020 ◽  
Vol 88 (8) ◽  
Author(s):  
Giorgio Mattiuz ◽  
Sabrina Nicolò ◽  
Alberto Antonelli ◽  
Tommaso Giani ◽  
Ilaria Baccani ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Lipid A ◽  
Nuclear Translocation ◽  
Interleukin 12 ◽  
Bacterial Survival ◽  
Necrosis Factor Alpha ◽  
Content Type ◽  
Human Macrophages ◽  
E Coli ◽  
The Impact

ABSTRACT MCR-1 is a plasmid-encoded phosphoethanolamine transferase able to modify the lipid A structure. It confers resistance to colistin and was isolated from human, animal, and environmental strains of Enterobacteriaceae, raising serious global health concerns. In this paper, we used recombinant mcr-1-expressing Escherichia coli to study the impact of MCR-1 products on E. coli-induced activation of inflammatory pathways in activated THP-1 cells, which was used as a model of human macrophages. We found that infection with recombinant mcr-1-expressing E. coli significantly modulated p38-MAPK and Jun N-terminal protein kinase (JNK) activation and pNF-κB nuclear translocation as well as the expression of genes for the relevant proinflammatory cytokines tumor necrosis factor alpha (TNF-α), interleukin-12 (IL-12), and IL-1β compared with mcr-1-negative strains. Caspase-1 activity and IL-1β secretion were significantly less activated by mcr-1-positive E. coli strains than the mcr-1-negative parental strain. Similar results were obtained with clinical isolates of mcr-1-positive E. coli, suggesting that, in addition to colistin resistance, the expression of mcr-1 allows the escape of early host innate defenses and may promote bacterial survival.

scholarly journals Characterization of human and mouse granulocyte-macrophage-colony-stimulating factors derived from Escherichia coli

1987 ◽  
Vol 247 (1) ◽  
pp. 195-199 ◽  
Author(s):  
J L Schrimsher ◽  
K Rose ◽  
M G Simona ◽  
P Wingfield
Keyword(s):  
Escherichia Coli ◽  
Amino Acid ◽  
Amino Acid Sequences ◽  
Natural Material ◽  
Animal Cells ◽  
Colony Stimulating Factors ◽  
E Coli ◽  
Macrophage Colony ◽  
The One ◽  
Human And Mouse

Human and mouse granulocyte-macrophage-colony-stimulating factors (hGM-CSF and mGM-CSF, respectively), isolated from Escherichia coli cells expressing the corresponding human and mouse genes, have been characterized. The observed properties of the proteins have been compared with those properties which can be deduced from the DNA sequence alone and the published properties of natural GM-CSFs. The purified E. coli-derived proteins were found to have the expected molecular masses, amino acid compositions and N- and C-terminal amino acid sequences. The finding of 70-90% unprocessed N-terminal methionine for both proteins is discussed. The four Cys residues were found to be involved in two intramolecular disulphide bonds, linking the first and third, and second and fourth Cys residues. This disulphide bond arrangement is probably the one existing in natural material, since, although not glycosylated, both E. coli-derived proteins showed biological activity (colony stimulating assay for hGM-CSF, and cell proliferation assay for mGM-CSF) comparable with that reported for the respective proteins purified from animal cells.

scholarly journals Conversion of Norepinephrine to 3,4-Dihdroxymandelic Acid in Escherichia coli Requires the QseBC Quorum-Sensing System and the FeaR Transcription Factor

2017 ◽  
Vol 200 (1) ◽  
Author(s):  
Sasikiran Pasupuleti ◽  
Nitesh Sule ◽  
Michael D. Manson ◽  
Arul Jayaraman
Keyword(s):  
Escherichia Coli ◽  
Transcription Factor ◽  
Quorum Sensing ◽  
Aromatic Amines ◽  
Response Regulator ◽  
Content Type ◽  
E Coli ◽  
Expression Of Genes ◽  
K 12 ◽  
Cognate Response Regulator

ABSTRACTThe detection of norepinephrine (NE) as a chemoattractant byEscherichia colistrain K-12 requires the combined action of the TynA monoamine oxidase and the FeaB aromatic aldehyde dehydrogenase. The role of these enzymes is to convert NE into 3,4-dihydroxymandelic acid (DHMA), which is a potent chemoattractant sensed by the Tsr chemoreceptor. These two enzymes must be induced by prior exposure to NE, and cells that are exposed to NE for the first time initially show minimal chemotaxis toward it. The induction of TynA and FeaB requires the QseC quorum-sensing histidine kinase, and the signaling cascade requires new protein synthesis. Here, we demonstrate that the cognate response regulator for QseC, the transcription factor QseB, is also required for induction. The related quorum-sensing kinase QseE appears not to be part of the signaling pathway, but its cognate response regulator, QseF, which is also a substrate for phosphotransfer from QseC, plays a nonessential role. The promoter of thefeaRgene, which encodes a transcription factor that has been shown to be essential for the expression oftynAandfeaB, has two predicted QseB-binding sites. One of these sites appears to be in an appropriate position to stimulate transcription from the P1promoter of thefeaRgene. This study unites two well-known pathways: one for expression of genes regulated by catecholamines (QseBC) and one for expression of genes required for metabolism of aromatic amines (FeaR, TynA, and FeaB). This cross talk allowsE. colito convert the host-derived and chemotactically inert NE into the potent bacterial chemoattractant DHMA.IMPORTANCEThe chemotaxis ofE. coliK-12 to norepinephrine (NE) requires the conversion of NE to 3,4-dihydroxymandleic acid (DHMA), and DHMA is both an attractant and inducer of virulence gene expression for a pathogenic enterohemorrhagicE. coli(EHEC) strain. The induction of virulence by DHMA and NE requires QseC. The results described here show that the cognate response regulator for QseC, QseB, is also required for conversion of NE into DHMA. Production of DHMA requires induction of a pathway involved in the metabolism of aromatic amines. Thus, the QseBC sensory system provides a direct link between virulence and chemotaxis, suggesting that chemotaxis to host signaling molecules may require that those molecules are first metabolized by bacterial enzymes to generate the actual chemoattractant.

scholarly journals Complete Genome Sequence of Escherichia coli Siphophage Snoke

2019 ◽  
Vol 8 (40) ◽  
Author(s):  
James E. Corban ◽  
Jacob Gramer ◽  
Russell Moreland ◽  
Mei Liu ◽  
Jolene Ramsey
Keyword(s):  
Escherichia Coli ◽  
Genome Sequence ◽  
Complete Genome Sequence ◽  
Complete Genome ◽  
Negative Bacterium ◽  
Protein Coding ◽  
Gram Negative ◽  
Content Type ◽  
E Coli ◽  
Protein Coding Genes

Escherichia coli is a Gram-negative bacterium often found in animal intestinal tracts. Here, we present the genome of the Guernseyvirinae-like E. coli 4s siphophage Snoke. The 44.4-kb genome contains 81 protein-coding genes, for which 33 functions were predicted. The capsid morphogenesis gene in Snoke contains a large intein.

scholarly journals The Effect of Various Concentrations of Ethanol Extract of the Leaves of Paederia foetida L. on the Growth of Escherichia Coli Bacteria

2021 ◽  
Vol 11 (6) ◽  
pp. 61-67
Author(s):  
Hertina Silaban
Keyword(s):  
Escherichia Coli ◽  
Ethanol Extract ◽  
Inhibition Zone ◽  
Positive Control ◽  
Negative Control ◽  
Rare Plants ◽  
E Coli ◽  
Paederia Foetida ◽  
Escherichia Coli Bacteria ◽  
The One

Bacterial infection of Escherichia coli (E. coli) as the cause of gastrointestinal disorders in humans has increased their prevalence. Treatment using natural ingredients can be a choice of therapy because of the minimal side effects. One of the rare plants believed by the community as an antibacterial is stinking vin’e known as the ‘leaf fart’. The purpose of this research is for knowing the activity of the ethanol extract of Paederia foetida L can affect the growth of E.coli. The serial diffusion disc method is being used as the antibacterial activity test. The concentration  of this extract are 10%, 20%, 40%, 80%, 100% with positive control (ciprofloxacin) and negative control (aqua dest). The inhibition zone diameter characterized the effect of Extract on bacterial growth were 6.16 mm of the concentration 10%, 6.667 mm of the concentration 20%, 7.10 mm of the concentration 40 %, 7.78 mm of the concentration 80%, and 10.03 mm of the concentration 100%. As for the negative control has no effect. The study stated that the higher concentration of antibacterial agent used, the greater the inhibition zone formed. Based on the result of the analysis of the data by using the One-Way ANOVA Test showed a probability value (p) = 0.000 or value (p) < 0.05, that H0 is rejected and H1 is accepted. The conclusion is that the Extract of stinking vin’e has an antibacterial effect on the growth of E.coli. Keywords: Antibacterial, E.coli, Extract of  Sembukan leaf

A Nursery Outbreak of Multiple-Aminoglycoside-Resistant Escherichia coli

1984 ◽  
Vol 5 (11) ◽  
pp. 519-524 ◽  
Author(s):  
Robert R Gaynes ◽  
Diane Simpson ◽  
Sally A. Reeves ◽  
Robert C. Noble ◽  
Clyde Thornsberry ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Neonatal Intensive Care ◽  
Control Measures ◽  
Effective Control ◽  
Blood Isolate ◽  
E Coli ◽  
Feeding Duration ◽  
Matched Case ◽  
Continuous Feeding ◽  
The One

AbstractIn the neonatal intensive care unit (NICU) of one hospital, 16 infants became colonized or infected with multiply-resistant Escherichia coli (MR-E.coli) over an 8-month period. Isolates were obtained from blood, urine, and sputum of three patients and from rectal surveillance cultures of 13 patients. The one patient with the blood isolate died. A matched case-control study identified continuous feeding (nine of 16 cases vs. one of 16 controls, p ≤ 0.001) and receipt of aminoglycosides (p ≤ 0.03) as risk factors. For case-babies not exposed to continuous feeding, duration of bolus feeding was significantly greater than for their controls (cases, 22 days; controls, 7 days; p ≤ 0.02). All 16 isolates were the same serotype and were resistant to amikacin, tobramycin, kanamycin, and gentamicin. The epidemiologic investigation suggested that MR-E. coli may have spread from person-to-person on the hands of personnel and that MR-E. coli persisted in the NICU for 8 months until effective control measures were instituted.

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