کد مقاله کد نشریه سال انتشار مقاله انگلیسی نسخه تمام متن
5131699 1491330 2017 10 صفحه PDF دانلود رایگان
عنوان انگلیسی مقاله ISI
Novel systematic detergent screening method for membrane proteins solubilization
موضوعات مرتبط
مهندسی و علوم پایه شیمی شیمی آنالیزی یا شیمی تجزیه
پیش نمایش صفحه اول مقاله
Novel systematic detergent screening method for membrane proteins solubilization
چکیده انگلیسی

Membrane proteins play crucial role in many cellular processes including cell adhesion, cell-cell communication, signal transduction and transport. To better understand the molecular basis of such central biological machines and in order to specifically study their biological and medical role, it is necessary to extract them from their membrane environment. To do so, it is challenging to find the best solubilization condition. Here we describe, a systematic screening method called BMSS (Biotinylated Membranes Solubilization & Separation) that allow screening 96 conditions at once. Streptavidine magnetic beads are used to separate solubilized proteins from remaining biotinylated membranes after solubilization. Relative quantification of dot blots help to select the best conditions to be confirmed by classical ultra-centrifugation and western blot. Classical detergents with different physical-chemical characteristics, novel calixarene based detergents and combination of both, were used for solubilization trials to obtain broad spectrum of conditions. Here, we show the application of BMSS to discover solubilization conditions of a GPCR target (MP-A) and a transporter (MP-B). The selected conditions allowed the solubilization and purification of non-aggregated and homogenous native membrane proteins A and B. Taken together, BMSS represent a rapid, reproducible and high throughput assessment of solubilization toward biochemical/functional characterization, biophysical screening and structural investigations of membrane proteins of high biological and medical relevance.

ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Analytical Biochemistry - Volume 517, 15 January 2017, Pages 40-49
نویسندگان
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