Substrate specificity of α‐humulene synthase from Zingiber zerumbet Smith and determination of kinetic constants by a spectrophotometric assay |
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| Authors: | Semra Alemdar Jan Christoph König Katja Seidel Andreas Kirschning Thomas Scheper Sascha Beutel |
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| Affiliation: | 1. Institute of Technical Chemistry, Leibniz Universit?t Hannover, Hannover, Germany;2. Institute of Organic Chemistry, Leibniz Universit?t Hannover, Hannover, Germany |
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| Abstract: | Terpene synthases are the key enzymes in terpene biosynthesis that provide a structurally complex and highly diverse product spectrum. A suitable and reliable analytical assay is indispensable to measure terpene synthase activity accurately and precisely. In this study, a malachite green assay (MG) was adapted to rapidly assay terpene synthase activity and was validated in comparison to an already established gas chromatography assay. A linear correlation between both assays was observed. Kinetic properties for the previously described sesquiterpene synthase α‐humulene synthase (HUM) from Zingiber zerumbet Smith were investigated for the bioconversion of the monoterpene precursors geranyl pyrophosphate (2E‐GPP) and neryl pyrophosphate (2Z‐NPP) as well as for the sesquiterpene precursor farnesyl pyrophosphate (2E,6E‐FPP). Also, gas chromatography mass spectrometry (GS‐MS) was carried out to identify the products of the bioconversion of (2E)‐GPP and (2Z)‐NPP. |
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| Keywords: | α ‐Humulene synthase Activity assay Malachite green Substrate specificity Terpene synthase |
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