کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
10843470 | 1069260 | 2007 | 4 صفحه PDF | دانلود رایگان |
عنوان انگلیسی مقاله ISI
Expression in Escherichia coli and in vitro refolding of the plant transcription factor Arabidopsis thaliana RGL3
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موضوعات مرتبط
علوم زیستی و بیوفناوری
بیوشیمی، ژنتیک و زیست شناسی مولکولی
زیست شیمی
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چکیده انگلیسی
Recombinant Arabidopsis thaliana (At) RGL-3, using two vectors pMAL-c2 and pET 21, was expressed as inclusion bodies in Escherichia coli under a range of temperature conditions. Only low levels (8-12% of total protein) of soluble protein were produced. The “soluble” fraction was shown by native PAGE to exist as soluble aggregates of RGL-3. A method was developed, consisting of induction of expression at various temperatures that yielded high levels of refoldable inclusion bodies using the pET vector. (At) RGL-3, as inclusion bodies, was solubilized in 8Â M urea and refolding was initiated by 20-fold direct dilution of denaturant. Under optimal conditions, 87% of the denatured protein of inclusion bodies was successfully re-natured. Refolding was monitored by “native” PAGE. Refolded RGL-3 was shown to be present as monomers and dimers. Attempts to further purify His-tagged RGL-3 using Ni/NTA chromatography resulted in the formation of higher polymers.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Protein Expression and Purification - Volume 53, Issue 2, June 2007, Pages 289-292
Journal: Protein Expression and Purification - Volume 53, Issue 2, June 2007, Pages 289-292
نویسندگان
Taha H. Al-Samarrai, Christopher A. Kirk, William T. Jones, Dawn Harvey, Xiaolin Sun,