کد مقاله کد نشریه سال انتشار مقاله انگلیسی نسخه تمام متن
8360091 1542327 2015 6 صفحه PDF دانلود رایگان
عنوان انگلیسی مقاله ISI
Expression, purification and crystallization of the (3R)-hydroxyacyl-ACP dehydratase HadAB complex from Mycobacterium tuberculosis
موضوعات مرتبط
علوم زیستی و بیوفناوری بیوشیمی، ژنتیک و زیست شناسی مولکولی زیست شیمی
پیش نمایش صفحه اول مقاله
Expression, purification and crystallization of the (3R)-hydroxyacyl-ACP dehydratase HadAB complex from Mycobacterium tuberculosis
چکیده انگلیسی
The (3R)-hydroxyacyl-ACP dehydratase HadAB, involved in the biosynthetic pathway for mycolic acid (MA) of Mycobacterium tuberculosis, catalyzes the third step in the fatty acid (FA) elongation cycle, which is an ideal and actual target for anti-tubercular agent. Though HadAB is predicted to be a member of the hotdog superfamily, it shares no sequence identity with typical hotdog fold isoenzyme FabZ. To characterize the significance of HadAB from the perspective of structural biology, large amount of pure HadAB complex is required for biochemical characterization and crystallization. Here, we used a unique expression and purification method. HadA and HadB were cloned separately and co-expressed in Escherichia coli. After GST affinity chromatography, two steps of anion exchange chromatography and gel filtration, the purity of the protein as estimated by SDS-PAGE was >95%. Using hanging-drop vapor-diffusion method, crystals were obtained and diffracted X-rays to 1.75 Å resolution. The crystal belongs to space group P41212, with unit-cell parameters a = b = 82.0 Å, c = 139.8 Å, α = β = γ = 90.0°.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Protein Expression and Purification - Volume 114, October 2015, Pages 115-120
نویسندگان
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