Hosoi Madoka, Ito Michiho, Yagura Toru, Adams Robert Phillip, Honda Gisho
Graduate School of Pharmaceutical Sciences, Kyoto University, Japan.
Biol Pharm Bull. 2004 Dec;27(12):1979-85. doi: 10.1248/bpb.27.1979.
cDNA cloning of a monoterpene synthase from Perilla frutescens whose steam-distilled oil contains 92.9% perillaketone, was performed by the PCR method using primers designed based on limonene synthase. The full-length nucleotide sequence of this cDNA consisted of 1978 bp including a 1827-bp translational region encoding a deduced protein of 608 amino acids, which was similar to that of limonene synthase from P. frutescens (85% identity). Functional expression of this clone in Escherichia coli yielded an active monoterpene synthase enzyme, which converted geranyl diphosphate into 53.8% myrcene, 20.9% sabinene, 19.8% linalool and 5.5% limonene. As for the extraction of reaction products, we performed SPME (solid phase micro extraction) as well as conventional solvent extraction, and compared these two extraction methods.
通过PCR方法，使用基于柠檬烯合酶设计的引物，对紫苏的单萜合酶进行了cDNA克隆，紫苏的水蒸气蒸馏油中含有92.9%的紫苏酮。该cDNA的全长核苷酸序列由1978 bp组成，包括一个1827 bp的翻译区域，编码一个推导的608个氨基酸的蛋白质，该蛋白质与紫苏柠檬烯合酶相似（同一性为85%）。该克隆在大肠杆菌中的功能表达产生了一种活性单萜合酶，它将香叶基二磷酸转化为53.8%的月桂烯、20.9%的桧烯、19.8%的芳樟醇和5.5%的柠檬烯。至于反应产物的提取，我们进行了固相微萃取（SPME）以及传统的溶剂萃取，并比较了这两种萃取方法。
