Development of fast and simple chromogenic methods for glucan phosphatases in-gel activity assays

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• 作者：Julieta B. Carrillo ; Diego F. Gomez-Casati ; Maria V. Busi ; ^{busi@cefobi-conicet.gov.ar} ; Mariana Martí ; n ; ^{martin@cefobi-conicet.gov.ar}
• 关键词：Glucan phosphatase ; In-gel assay ; Chromogenic substrate ; Native-PAGE
• 刊名：Analytical Biochemistry
• 出版年：2017
• 出版时间：15 January 2017
• 年：2017
• 卷：517
• 期：Complete
• 页码：36-39
• 全文大小：626 K
• 卷排序：517

文摘

Glucan phosphatases are essential for normal starch degradation in plants and glycogen metabolism in mammals. Here we develop two chromogenic methods for the detection of glucan phosphatase activity in situ after non denaturing poliacrylamide gel electrophoresis; one method uses pNPP and the second one applies BCIP/NBT. The assays are sensitive, fast, simple, reliable and cost-effective preventing the use of radioactive or fluorogenic compounds.Taking advantage of an efficient separation method combined with the reported assays it is possible to obtain information about oligomeric state of the active enzymes as well as to simultaneously detect glucan substrate binding and phosphatase activity.