کد مقاله کد نشریه سال انتشار مقاله انگلیسی نسخه تمام متن
6488125 1416358 2018 6 صفحه PDF دانلود رایگان
عنوان انگلیسی مقاله ISI
A simplified method for active-site titration of lipases immobilised on hydrophobic supports
ترجمه فارسی عنوان
یک روش ساده برای تیتراسیون فعال سایت های لیپاز که بر روی حمایت های هیدروفوب قرار گرفته اند
موضوعات مرتبط
مهندسی و علوم پایه مهندسی شیمی بیو مهندسی (مهندسی زیستی)
چکیده انگلیسی
The aim of this work was to develop a simple and accurate protocol to measure the functional active site concentration of lipases immobilised on highly hydrophobic supports. We used the potent lipase inhibitor methyl 4-methylumbelliferyl hexylphosphonate to titrate the active sites of Candida rugosa lipase (CrL) bound to three highly hydrophobic supports: octadecyl methacrylate (C18), divinylbenzene crosslinked methacrylate (DVB) and styrene. The method uses correction curves to take into account the binding of the fluorophore (4-methylumbelliferone, 4-MU) by the support materials. We showed that the uptake of the detection agent by the three supports is not linear relative to the weight of the resin, and that the uptake occurs in an equilibrium that is independent of the total fluorophore concentration. Furthermore, the percentage of bound fluorophore varied among the supports, with 50 mg of C18 and styrene resins binding approximately 64 and 94%, respectively. When the uptake of 4-MU was calculated and corrected for, the total 4-MU released via inhibition (i.e. the concentration of functional lipase active sites) could be determined via a linear relationship between immobilised lipase weight and total inhibition. It was found that the functional active site concentration of immobilised CrL varied greatly among different hydrophobic supports, with 56% for C18, compared with 14% for DVB. The described method is a simple and robust approach to measuring functional active site concentration in immobilised lipase samples.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Enzyme and Microbial Technology - Volume 113, June 2018, Pages 18-23
نویسندگان
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