کد مقاله کد نشریه سال انتشار مقاله انگلیسی نسخه تمام متن
2020938 1069218 2010 6 صفحه PDF دانلود رایگان
عنوان انگلیسی مقاله ISI
Expression of the functional carbohydrate-binding module (CBM) of human laforin
موضوعات مرتبط
علوم زیستی و بیوفناوری بیوشیمی، ژنتیک و زیست شناسی مولکولی زیست شیمی
پیش نمایش صفحه اول مقاله
Expression of the functional carbohydrate-binding module (CBM) of human laforin
چکیده انگلیسی

Laforin is a human protein associated with the glycogen metabolism, composed of two structurally and functionally independent domains: a phosphatase catalytic domain and a substrate-binding module with glycogen and starch affinity. The main goal of this work is the development of a methodology for the expression of the so far poorly characterized carbohydrate-binding module (CBM) of laforin, allowing its study and development of biomedical applications. The laforin’s CBM sequence was originally cloned by PCR from a human muscle cDNA library. The recombinant protein, containing laforin’s CBM fused to an Arg-Gly-Asp sequence (RGD), was cloned and expressed using vector pET29a and recovered as inclusion bodies (IBs). Refolding of the IBs allowed the purification of soluble, dimeric and functional protein, according to adsorption assays using starch and glycogen. Several other experimental approaches, using both bacteria and yeast, were unsuccessfully tested, pointing towards the difficulties in producing the heterologous protein. Indeed, this is the first work reporting the production of the functional CBM from human laforin.

ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Protein Expression and Purification - Volume 74, Issue 2, December 2010, Pages 169–174
نویسندگان
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