A HIGH-THROUGHPUT IN-VITRO ASSAY FOR FUNGAL (1,3)BETA-GLUCAN SYNTHASE INHIBITORS

被引：12

作者：

TAFT, CS

ENDERLIN, CS

SELITRENNIKOFF, CP

机构：

[1] Mycotox, Inc., Denver, CO 80262

来源：

JOURNAL OF ANTIBIOTICS | 1994年 / 47卷 / 09期

关键词：

D O I：

10.7164/antibiotics.47.1001

中图分类号：

Q81 [生物工程学（生物技术）]; Q93 [微生物学];

学科分类号：

071005 ; 0836 ; 090102 ; 100705 ;

摘要：

An in vitro assay for (1,3)beta-glucan synthase activity from the filamentous ascomycete Neurospora crassa, suitable for use as a high throughput screen for enzyme inhibitors is described. Samples were added to 25 mu l reaction mixtures in 96-well V-bottom microtiter plates and plates incubated at 22 degrees C. Reactions were terminated by the addition of TCA and the contents transferred to a Milliblot D apparatus containing Inotech 201-A glass fiber filters. Filters were washed to remove unincorporated substrate and the amount of C-14-labeled (1,3)beta-glucan formed by each reaction was quantitated using a Phosphorlmager. As little as 50 ng of an inhibitor with a Ki of 4 mu M was detected by this assay. This assay is rapid and cost effective, permitting its use as a screen to detect compounds that inhibit (1,3)beta-glucan synthase activity.

引用

页码：1001 / 1009

页数：9

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