Department of Pharmacognosy, Graduate School of Pharmaceutical Sciences, Kyoto University, 46-29 Yoshidashimoadachi-cho, Sakyo-ku, Kyoto, 606-8501, Japan.
J Nat Med. 2021 Jun;75(3):577-589. doi: 10.1007/s11418-021-01503-3. Epub 2021 Mar 29.
Phenylpropanoid volatile components are found in various plants and are useful in medicines, health foods, and fragrances. While the pharmacological actions and toxicities of these compounds have been investigated, there are few reports of the cloning of genes that encode biosynthetic enzymes involved in substituent formation at diverse positions and numbers. Previously, using the expressed sequence tag (EST) libraries of pure perilla strains that have been maintained for over 30 years for their oil type, we characterized the P450 enzyme that produces an intermediate for dillapiole by adding a hydroxy group to myristicin. In this study, we selected a P450 enzyme involved in nothoapiole biosynthesis from the EST library. Heterologous expression of this enzyme in yeast showed that it is a hydroxylase that synthesizes an intermediate to produce nothoapiole from apiole and dillapiole. The enzyme has high amino acid sequence similarity with a previously cloned enzyme and is categorized into the CYP71D subfamily. Furthermore, we investigated the presence or absence of essential oil components and intermediates believed to be involved in nothoapiole biosynthesis by component analysis of perilla essential oil using GC-MS to help elucidate the biosynthetic pathway of nothoapiole. Only a small number of plant species contain nothoapiole as their principal component and thus few studies have reported the biosynthetic genes involved or the drug efficacy and toxicity of nothoapiole. The present study will aid in understanding the biosynthesis of phenylpropanoid volatile compounds, thereby contributing to further research on potentially useful compounds such as nothoapiole.
苯丙素类挥发性成分存在于各种植物中，在医药、保健食品和香料中都有应用。虽然这些化合物的药理作用和毒性已经得到了研究，但关于编码不同位置和数量取代基形成的生物合成酶的基因克隆的报道却很少。之前，我们利用经过 30 多年油型选育的纯紫苏品系的表达序列标签（EST）文库，对 P450 酶进行了研究，该 P450 酶通过向肉豆蔻醚中添加一个羟基来生成紫穗槐烯酮的中间产物。在这项研究中，我们从 EST 文库中选择了一种参与白花前胡醇生物合成的 P450 酶。该酶在酵母中的异源表达表明，它是一种羟化酶，能够从茴香脑和紫穗槐烯酮合成白花前胡醇的中间产物。该酶与之前克隆的酶具有高度的氨基酸序列相似性，属于 CYP71D 亚家族。此外，我们通过 GC-MS 对紫苏精油的成分分析，研究了认为与白花前胡醇生物合成有关的精油成分和中间产物的存在与否，以帮助阐明白花前胡醇的生物合成途径。只有少数植物物种将白花前胡醇作为其主要成分，因此很少有研究报道与白花前胡醇生物合成有关的基因或白花前胡醇的药效和毒性。本研究将有助于理解苯丙素类挥发性化合物的生物合成，从而为进一步研究可能有用的化合物如白花前胡醇提供帮助。
