scholarly journals In VitroActivation and Inhibition of Recombinant EGFR Tyrosine Kinase Expressed inEscherichia coli

2013 ◽  
Vol 2013 ◽  
pp. 1-5 ◽  
Author(s):  
Jihene Elloumi-Mseddi ◽  
Karim Jellali ◽  
Sami Aifa
Keyword(s):  
Tyrosine Kinase ◽  
Kinase Activity ◽  
Growth Factor Receptor ◽  
Kinase Domain ◽  
Tyrosine Kinase Domain ◽  
Glutathione S Transferase ◽  
Amino Acid Peptide ◽  
Juxtamembrane Region ◽  
Exogenous Substrate ◽  
Epidermal Growth

The present work concerns the heterologous expression of the intracellular domain harbouring the tyrosine kinase activity of the epidermal growth factor receptor (EGFR). Protein expression was improved thanks to the deletion of a 13-amino acid peptide of the juxtamembrane region (JM). The recombinant proteins were produced as a glutathione S-transferase (GST) fusion inEscherichia coli, and the solubilisation was performed by sarkosyl addition during extraction. The produced proteins spontaneously dimerize allowing the activation of the tyrosine kinase domain in the presence of[γ-32P]ATP. The activity assay has revealed the autophosphorylation of EGFR proteins which was decreased in the presence of genistein. Our system could facilitate the screening of EGFR inhibitors without the need of adding an exogenous substrate.

scholarly journals Protein tyrosine kinase activities of the epidermal growth factor receptor and ErbB proteins: correlation of oncogenic activation with altered kinetics

1992 ◽  
Vol 12 (5) ◽  
pp. 2010-2016
Author(s):  
N Nair ◽  
R J Davis ◽  
H L Robinson
Keyword(s):  
Epidermal Growth Factor Receptor ◽  
Tyrosine Kinase ◽  
Protein Tyrosine Kinase ◽  
Kinase Activity ◽  
Growth Factor Receptor ◽  
Tyrosine Kinase Activity ◽  
Protein Tyrosine ◽  
Exogenous Substrate ◽  
Epidermal Growth ◽  
Kinetics Of

We have compared the protein tyrosine kinase activities of the chicken epidermal growth factor receptor (chEGFR) and three ErbB proteins to learn whether cancer-activating mutations affect the kinetics of kinase activity. In immune complex assays performed in the presence of 15 mM Mn2+, ErbB proteins and the chEGFR exhibited highly reproducible tyrosine kinase activity. Under these conditions, the ErbB and chEGFR proteins had similar apparent Km [Km(app)] values for ATP. The ErbB proteins appeared to be activated, as they had at least 3-fold-higher relative Vmax(app) for autophosphorylation and approximately 2-fold higher relative Vmax(app) for the phosphorylation of the exogenous substrate TK6 (a bacterially expressed fusion protein containing the C-terminal domain of the human EGFR). The ErbB kinases had both higher Km(app) and higher Vmax(app) for the phosphorylation of the exogenous substrate TK6 than did the chEGFR. The ratios of the Vmax(app) to the Km(app) for TK6 phosphorylation suggested that the ErbB proteins had lower catalytic efficiencies for the exogenous substrate than did the chEGFR. The three tested ErbB proteins had cytoplasmic domain mutations that conferred distinctive disease potentials. These mutations did not affect the kinetics for the phosphorylation of the exogenous substrate TK6. Two of the ErbB proteins contained all of the sites used for autophosphorylation. In these, a mutation that broadened oncogenic potential to endothelial cells caused an additional increase in Vmax(app) for autophosphorylation. Thus, mutations that change the EGFR into an ErbB oncogene cause multiple changes in the kinetics of protein tyrosine kinase activity.

scholarly journals Large-cell neuroendocrine carcinoma of lung with epidermal growth factor receptor (EGFR) gene mutation and co-expression of adenocarcinoma markers: a case report and review of the literature

2013 ◽  
Vol 8 ◽  
Author(s):  
Yasuhiro Sakai ◽  
Takashi Yamasaki ◽  
Yoshito Kusakabe ◽  
Daisuke Kasai ◽  
Yoshikazu Kotani ◽  
...  
Keyword(s):  
Tyrosine Kinase ◽  
Neuroendocrine Carcinoma ◽  
Growth Factor Receptor ◽  
Large Cell ◽  
Kinase Domain ◽  
Large Cell Neuroendocrine Carcinoma ◽  
Tyrosine Kinase Domain ◽  
High Grade ◽  
Epidermal Growth ◽  
L858r Mutation

Purpose: A high rate of response to treatment with epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI) has been observed in certain patients (women, of East Asian ethnicity, with non-smoking history and adenocarcinoma histology) with mutations in exons 18 to 21 of the tyrosine kinase domain of EGFR. Some cases of high-grade neuroendocrine carcinoma of the lung harboring mutations have been sporadically reported. Methods: We describe the case of a 78-year-old woman with large-cell neuroendocrine carcinoma of the lung, with mutation in exon 21 L858R and co-expression of adenocarcinoma markers. Results: A mass (3.0 cm in diameter) was identified in the inferior lobe of the left lung, accompanied by metastases into ipsilateral mediastinal lymph nodes and elevations of serum pro-gastrin-releasing peptide and carcinoembryonic antigen. Initial transbronchial brushing cytology suggested high-grade neuroendocrine carcinoma favoring small-cell carcinoma in poorly smeared and degenerated preparations, and revealed exon 21 L858R mutation. Re-enlargement of the cancer and bone metastases was observed after chemotherapy, and further testing suggested large-cell neuroendocrine carcinoma with immunoreactivity to markers of primary lung adenocarcinoma and L858R mutation. High-grade neuroendocrine carcinoma with mutations in the tyrosine kinase domain of EGFR may be associated with adenocarcinoma, as reviewed from the literature and may also apply to our case. Conclusions: EGFR-TKI could provide better quality of life and survival in patients with advanced or relapsed high-grade neuroendocrine carcinoma with EGFR gene mutations. Further studies in this respect are warranted.

scholarly journals Use of Marine biotoxins to modulate the tyrosine kinase domain of the human epidermal growth factor receptor

2021 ◽  
Author(s):  
Charli Deepak Arulanandam ◽  
Ramesh Dharmara ◽  
Prathiviraj Ragothaman ◽  
Samuel Gnana Prakash Vincent
Keyword(s):  
Epidermal Growth Factor Receptor ◽  
Molecular Docking ◽  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Tyrosine Kinase ◽  
Growth Factor Receptor ◽  
Kinase Domain ◽  
Tyrosine Kinase Domain ◽  
Marine Biotoxins ◽  
Epidermal Growth

Inappropriate activation of the Epidermal growth factor receptor (EGFR) group of kinases has been identified in a variety of tumour cells, either due to mutation or overexpression. Although the tumour is a fatal disease, significant therapy discoveries have lately been made. The human EGFR and this family of kinases have emerged as promising targets for cancer therapy. In this molecular docking study, Natural marine toxins are employed to regulate the activity of the human EGFR tyrosine kinase domain (EGFRtkd) in the molecular docking investigation (PDB ID5JEB). Marine biotoxins can cause neurological, gastrointestinal, and cardiovascular problems, as well as severe mortality and long-term morbidity in some situations. Because there is no antidote for any of the natural marine poisons, supportive care is the mainstay of treatment. Paralytic shellfish poisoning, in particular, and puffer fish poisoning, in particular, can result in death within hours of exposure to the poisons and may require immediate medical intervention. However, this research found that marine biotoxins can modulate EGFRtkd. Furthermore, homoyessotoxin was anticipated to be an EGFRtkd modulator with a binding affinity as -9.584 kcal/mol. To employ the homoyessotoxin in tumour therapies, further knowledge of natural marine biotoxins and further toxicological research is required.

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