کد مقاله کد نشریه سال انتشار مقاله انگلیسی نسخه تمام متن
2058699 1543969 2014 10 صفحه PDF دانلود رایگان
عنوان انگلیسی مقاله ISI
Comparative analysis and validation of the malachite green assay for the high throughput biochemical characterization of terpene synthases
ترجمه فارسی عنوان
تجزیه و تحلیل تطبیقی ​​و اعتبار سنجی از آزمون سبز مالاشیوت برای مشخصه بیوشیمیایی بالا سنتز ترپن
موضوعات مرتبط
علوم زیستی و بیوفناوری بیوشیمی، ژنتیک و زیست شناسی مولکولی بیوشیمی، ژنتیک و زیست شناسی مولکولی (عمومی)
چکیده انگلیسی

Terpenes are the largest group of natural products with important and diverse biological roles, while of tremendous economic value as fragrances, flavours and pharmaceutical agents. Class-I terpene synthases (TPSs), the dominant type of TPS enzymes, catalyze the conversion of prenyl diphosphates to often structurally diverse bioactive terpene hydrocarbons, and inorganic pyrophosphate (PPi). To measure their kinetic properties, current bio-analytical methods typically rely on the direct detection of hydrocarbon products by radioactivity measurements or gas chromatography–mass spectrometry (GC–MS).In this study we employed an established, rapid colorimetric assay, the pyrophosphate/malachite green assay (MG), as an alternative means for the biochemical characterization of class I TPSs activity.
• We describe the adaptation of the MG assay for turnover and catalytic efficiency measurements of TPSs.
• We validate the method by direct comparison with established assays. The agreement of kcat/KM among methods makes this adaptation optimal for rapid evaluation of TPSs.
• We demonstrate the application of the MG assay for the high-throughput screening of TPS gene libraries.

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ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: MethodsX - Volume 1, 2014, Pages 187–196
نویسندگان
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