Centro de Ciências do Mar, Universidade do Algarve, Campus de Gambelas, 8005-139 Faro, Portugal.
Centre for Mediterranean Bioresources and Food (MeditBio), Faculdade de Ciências e Tecnologia, Universidade do Algarve, Campus de Gambelas, 8005-139 Faro, Portugal.
J Plant Physiol. 2017 Nov;218:35-44. doi: 10.1016/j.jplph.2017.07.013. Epub 2017 Jul 21.
The essential oil of Thymus albicans Hoffmanns. & Link, a native shrub from the Iberian Peninsula, is mainly composed of monoterpenes. In this study, a 1,8-cineole synthase was isolated from the 1,8-cineole chemotype. A partial sequence that lacked the complete plastid transit peptide but contained an extended C-terminal when compared to other related terpene synthases was generated by PCR and Rapid Amplification of cDNA Ends (RACE). The predicted mature polypeptide was 593 amino acids in length and shared 78% and 77% sequence similarity with the homologue 1,8-cineole synthase from Rosmarinus officinalis and Salvia officinalis, respectively. The putative protein possessed the characteristic conserved motifs of plant monoterpene synthases including the RRxW and DDxxD motifs and phylogenetic analysis indicated that the amplified 1,8-cineole synthase bears greater sequence similarity with other 1,8-cineole synthases from Lamiaceae family relative to the terpene synthases from the genus Thymus. Functional expression of the recombinant protein in Escherichia coli revealed that in the presence of geranyl diphosphate (GPP) 1,8-cineole was the major product but that its production was too low for robust quantification. Other minor conversion products included α-pinene, β-pinene, sabinene and β-myrcene suggesting the isolated 1,8-cineole synthase may be a multi-product enzyme. To our knowledge, this is the first report of a functionally characterized monoterpene synthase from Thymus albicans.
从伊比利亚半岛土生土长的灌木百里香（Thymus albicans Hoffmanns. & Link）中提取的精油主要由单萜烯组成。本研究从 1,8-桉叶油醇化学型中分离出 1,8-桉叶醇合酶。通过 PCR 和快速扩增 cDNA 末端 (RACE) 生成了一个部分序列，该序列与其他相关萜烯合酶相比缺少完整的质体转运肽，但包含扩展的 C 端。预测的成熟多肽长 593 个氨基酸，与迷迭香（Rosmarinus officinalis）和鼠尾草（Salvia officinalis）的同源 1,8-桉叶醇合酶分别具有 78%和 77%的序列相似性。该假定蛋白具有植物单萜烯合酶的特征保守基序，包括 RxW 和 DDxxD 基序，系统发育分析表明，扩增的 1,8-桉叶醇合酶与唇形科家族的其他 1,8-桉叶醇合酶具有更大的序列相似性，而与百里香属的萜烯合酶相比则较低。该重组蛋白在大肠杆菌中的功能表达表明，在香叶基二磷酸（GPP）存在的情况下，1,8-桉叶醇是主要产物，但产量太低，无法进行稳健的定量。其他次要转化产物包括α-蒎烯、β-蒎烯、月桂烯和β-罗勒烯，表明分离的 1,8-桉叶醇合酶可能是一种多产物酶。据我们所知，这是首次从百里香属中鉴定出具有功能特征的单萜烯合酶。
