کد مقاله کد نشریه سال انتشار مقاله انگلیسی نسخه تمام متن
8359555 1542297 2018 13 صفحه PDF دانلود رایگان
عنوان انگلیسی مقاله ISI
Expression and purification of the human epidermal growth factor receptor extracellular domain
ترجمه فارسی عنوان
بیان و پاکسازی دامنه خارج سلولی گیرنده عامل اپیدرمی انسانی
کلمات کلیدی
گیرنده فاکتور رشد اپیدرمال، پروتئین های پستانداران، پروتئین مجدد
موضوعات مرتبط
علوم زیستی و بیوفناوری بیوشیمی، ژنتیک و زیست شناسی مولکولی زیست شیمی
چکیده انگلیسی
In the fields of drug discovery and protein science, small quantities of proteins are always needed to investigate or validate protein-protein (or protein-small molecule) interactions. Traditional transient or stable expression method to obtain recombinant proteins in eukaryotic systems can be laborious and time-consuming, especially when multiple protein variants are required. Here, we present a fast and convenient method for obtaining small quantities of recombinant human epidermal growth factor receptor (rhEGFR) ectodomain protein, which could be efficiently extended to the expression of other eukaryotic proteins. Human EGFR ectodomain gene was inserted into the plasmid pBMN-GFP and recombinant plasmid was transfected into HEK 293 T cells. In the presence of hygromycin, cells with the integrated human EGFR ectodomain gene were selected and proliferated. rhEGFR ectodomain in cell culture supernatant was purified using serial connected diethylaminoethyl Sepharose column and Ni-NTA Sepharose column. Purity of the final purified rhEGFR ectodomain was over 95% according to SDS-PAGE analysis. Moreover, the purified target protein was biological active via measuring the affinity between the rhEGFR ectodomain and rhEGF. Our method could greatly facilitate research in the areas of protein science, protein structural biology, and drug discovery.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Protein Expression and Purification - Volume 144, April 2018, Pages 33-38
نویسندگان
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