TY - JOUR

T1 - Substrate specificity of α-humulene synthase from Zingiber zerumbet Smith and determination of kinetic constants by a spectrophotometric assay

AU - Alemdar, Semra

AU - König, Jan Christoph

AU - Seidel, Katja

AU - Kirschning, Andreas

AU - Scheper, Thomas

AU - Beutel, Sascha

N1 - Funding information: This study was funded by the European Regional Development Fund (EFRE): Innovation Network “Refinement of plant resources” (ZW-8-80130940). Special thanks to Daniel Sandner and Dr. Ulrich Krings (both Institute of Food Chemistry, Leibniz Universität Hannover) for performing GS-MS measurements.

PY - 2018/9/3

Y1 - 2018/9/3

N2 - 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.

AB - 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.

KW - Activity assay

KW - Malachite green

KW - Substrate specificity

KW - Terpene synthase

KW - α-Humulene synthase

UR - http://www.scopus.com/inward/record.url?scp=85050392314&partnerID=8YFLogxK

U2 - 10.1002/elsc.201800019

DO - 10.1002/elsc.201800019

M3 - Article

C2 - 32624945

AN - SCOPUS:85050392314

VL - 18

SP - 654

EP - 658

JO - Engineering in life sciences

JF - Engineering in life sciences

SN - 1618-0240

IS - 9

ER -