کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
8359744 | 1542321 | 2016 | 9 صفحه PDF | دانلود رایگان |
عنوان انگلیسی مقاله ISI
Expression, purification and characterization of Plasmodium falciparum vacuolar protein sorting 29
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کلمات کلیدی
DLSRPMI-1640Roswell Park Memorial Institute-1640matrix-assisted laser desorption/ionization time-of-flight mass spectrometry - اسپکترومتر جرمی زمان یخ زدن لیزر جذب / یونیزاسیون ماتریسProtein expression and purification - بیان و پاکسازی پروتئینCloning - تاگسازی یا شبیه سازیcircular dichroism - رنگ تابی دورانیMALDI ToF MS - مالدی تاف MSDynamic Light Scattering - پراکندگی نور دینامیکیPlasmodium falciparum - پلاسمودیوم فالسیپاروم
موضوعات مرتبط
علوم زیستی و بیوفناوری
بیوشیمی، ژنتیک و زیست شناسی مولکولی
زیست شیمی
پیش نمایش صفحه اول مقاله
چکیده انگلیسی
Translocation of various proteins to the subcellular organelles is an essential mechanism to regulate the metabolic pathways and often vacuolar protein sorting (VPS) proteins are involved in this transportation. Plasmodium falciparum VPS29 (PfVPS29) is predicted to be a functional component in the assembly of the retromer complex; however, so far detailed characterization of PfVPS29 in its native form is not yet done. We report the successful expression and purification of tag-free recombinant PfVPS29 with a yield of 5.6 mg from 1 L of Escherichia coli culture. PfVPS29 was purified by combined anion-exchange and size exclusion chromatography. The protein showed a single band in SDS-PAGE and it exhibited molecular mass of 21.7 kDa as measured by MALDI-TOF mass spectrometry. Secondary structure was elucidated by circular dichroism spectroscopy. It was found to be a monomeric protein in solution as evident from dynamic light scattering studies, chemical cross-linking experiments and size exclusion chromatography. Subsequently, polyclonal anti-PfVPS29 antibody was generated and used for evaluating protein expression by western blot and following subcellular localization in P. falciparum by confocal immunofluoroscence microscopy. PfVPS29 was found to be located in cytoplasm and expressed from early trophozoite to schizont stages with maximum expression in trophozoite stage. This study provides purification, biophysical characterization and subcellular localization of PfVPS29 in different asexual stages of P. falciparum.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Protein Expression and Purification - Volume 120, April 2016, Pages 7-15
Journal: Protein Expression and Purification - Volume 120, April 2016, Pages 7-15
نویسندگان
Mohd. Shameel Iqbal, Asim Azhar Siddiqui, Athar Alam, Manish Goyal, Chinmoy Banerjee, Souvik Sarkar, Somnath Mazumder, Rudranil De, Shiladitya Nag, Shubhra Jyoti Saha, Uday Bandyopadhyay,