کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
11007552 | 1538379 | 2018 | 36 صفحه PDF | دانلود رایگان |
عنوان انگلیسی مقاله ISI
Elucidating paramylon and other carbohydrate metabolism in Euglena gracilis: Kinetic characterization, structure and cellular localization of UDP-glucose pyrophosphorylase
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کلمات کلیدی
موضوعات مرتبط
علوم زیستی و بیوفناوری
بیوشیمی، ژنتیک و زیست شناسی مولکولی
زیست شیمی
پیش نمایش صفحه اول مقاله
چکیده انگلیسی
Many oligo and polysaccharides (including paramylon) are critical in the Euglena gracilis life-cycle and they are synthesized by glycosyl transferases using UDP-glucose as a substrate. Herein, we report the molecular cloning of a gene putatively coding for a UDP-glucose pyrophosphorylase (EgrUDP-GlcPPase) in E. gracilis. After heterologous expression of the gene in Escherichia coli, the recombinant enzyme was characterized structural and functionally. Highly purified EgrUDP-GlcPPase exhibited a monomeric structure, able to catalyze synthesis of UDP-glucose with a Vmax of 3350 U.mgâ1. Glucose-1P and UTP were the preferred substrates, although the enzyme also used (with lower catalytic efficiency) TTP, galactose-1P and mannose-1P. Oxidation by hydrogen peroxide inactivated the enzyme, an effect reversed by reduction with dithiothreitol or thioredoxin. The redox process would involve sulfenic acid formation, since no pair of the 7 cysteine residues is close enough in the 3D structure of the protein to form a disulfide bridge. Electrophoresis studies suggest that, after oxidation, the enzyme arranges in many enzymatically inactive structural conformations; which were also detected in vivo. Finally, confocal fluorescence microscopy provided evidence for a cytosolic (mainly in the flagellum) localization of the enzyme.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Biochimie - Volume 154, November 2018, Pages 176-186
Journal: Biochimie - Volume 154, November 2018, Pages 176-186
نویسندگان
Robertino J. Muchut, Rodrigo D. Calloni, Fernando E. Herrera, Sergio A. Garay, Diego G. Arias, Alberto A. Iglesias, Sergio A. Guerrero,