کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
9894300 | 1542354 | 2005 | 5 صفحه PDF | دانلود رایگان |
عنوان انگلیسی مقاله ISI
An alternative use of basic pGEX vectors for producing both N- and C-terminal fusion proteins for production and affinity purification of antibodies
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کلمات کلیدی
موضوعات مرتبط
علوم زیستی و بیوفناوری
بیوشیمی، ژنتیک و زیست شناسی مولکولی
زیست شیمی
پیش نمایش صفحه اول مقاله
چکیده انگلیسی
Glutathione S-transferase (GST) fusion proteins are widely used in protein production for pure immunogens, protein-protein, and DNA-protein interaction studies. Using basic pGEX vectors, foreign DNA is introduced to the C-terminus of the GST gene and the produced fusion proteins are C-terminally orientated. However, because the orientation of foreign polypeptides may have a very important role in the correct folding of the produced polypeptides, N-terminal fusion proteins are needed to express especially the N-terminus of the foreign polypeptide. Here, we introduce a novel use of the basic pGEX vectors for the production of N-terminal fusion proteins. In this procedure, PCR generated DNA fragments were cloned into the N-terminus of the GST gene in a unique EcoNI site located down-stream of the ATG initiation codon. The N-terminal fusion proteins were expressed in high quantities, easily solubilized, and affinity purified using our modification of current purification protocols. We also introduce here a new modification of the affinity purification of antibodies using covalently crosslinked GST and fusion proteins to glutathione-agarose beads. Our procedure was tested successfully for producing antibodies against both N- and C-terminus of the luteinizing hormone/chorionic gonadotropin receptor.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Protein Expression and Purification - Volume 40, Issue 2, April 2005, Pages 287-291
Journal: Protein Expression and Purification - Volume 40, Issue 2, April 2005, Pages 287-291
نویسندگان
Jyrki T. Aatsinki, Hannu J. Rajaniemi,