Taura F, Morimoto S, Shoyama Y
Faculty of Pharmaceutical Sciences, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812, Japan.
J Biol Chem. 1996 Jul 19;271(29):17411-6. doi: 10.1074/jbc.271.29.17411.
We identified a unique enzyme that catalyzes the oxidocyclization of cannabigerolic acid to cannabidiolic acid (CBDA) in Cannabis sativa L. (CBDA strain). The enzyme, named CBDA synthase, was purified to apparent homogeneity by a four-step procedure: ammonium sulfate precipitation followed by chromatography on DEAE-cellulose, phenyl-Sepharose CL-4B, and hydroxylapatite. The active enzyme consists of a single polypeptide with a molecular mass of 74 kDa and a pI of 6.1. The NH2-terminal amino acid sequence of CBDA synthase is similar to that of Delta1-tetrahydrocannabinolic-acid synthase. CBDA synthase does not require coenzymes, molecular oxygen, hydrogen peroxide, and metal ion cofactors for the oxidocyclization reaction. These results indicate that CBDA synthase is neither an oxygenase nor a peroxidase and that the enzymatic cyclization does not proceed via oxygenated intermediates. CBDA synthase catalyzes the formation of CBDA from cannabinerolic acid as well as cannabigerolic acid, although the kcat for the former (0.03 s-1) is lower than that for the latter (0.19 s-1). Therefore, we conclude that CBDA is predominantly biosynthesized from cannabigerolic acid rather than cannabinerolic acid.
我们在大麻（CBDA 品系）中鉴定出一种独特的酶，它催化大麻二酚酸氧化环化生成大麻二酚（CBDA）。这种酶被命名为 CBDA 合酶，通过以下四步程序纯化至表观均一：硫酸铵沉淀，随后依次在 DEAE - 纤维素、苯基 - 琼脂糖 CL - 4B 和羟基磷灰石上进行层析。活性酶由一条单多肽链组成，分子量为 74 kDa，pI 为 6.1。CBDA 合酶的 NH2 末端氨基酸序列与 Δ1 - 四氢大麻酚酸合酶的相似。CBDA 合酶在氧化环化反应中不需要辅酶、分子氧、过氧化氢和金属离子辅因子。这些结果表明，CBDA 合酶既不是加氧酶也不是过氧化物酶，并且酶促环化反应不通过氧化中间体进行。CBDA 合酶催化从大麻萜酚酸以及大麻二酚酸形成 CBDA，尽管前者的 kcat（0.03 s-1）低于后者（0.19 s-1）。因此，我们得出结论，CBDA 主要是由大麻二酚酸而非大麻萜酚酸生物合成的。
