Purification OF THCA synthase from Cannabis callus

从大麻愈伤组织中纯化 THCA 合酶

• 批准号: 05671830
• 负责人: MORIMOTO Satoshi
• 金额: $ 1.34万
• 依托单位: KYUSHU UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (C)
• 财政年份: 1993
• 资助国家: 日本
• 起止时间: 1993 至 1994
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-05671830/
• 关键词: Cannabis sativa; Cannabinoid; Tetrahydrocannabinolic acid; Tetrahydrocannabinolic acid synthase; テトラハイドロカンナビノリックアシド; 生合成; Cannabis

The occurrence of DELTA^1-tetrahydrocannabinolic acid synthase was confirmed in the fresh leaf buds of marihuana (Cannabis sativa L.). The enzyme, assayd with cannabigerolic acid, was purified approximately 300-fold by chromatography on diethylaminoethyl-cellulose, phenyl Sepharose CL-4B and hydroxylapatite. The purified protein migrated during sodium dodecylsulfate-polyacrylamide gel electrophoresis (SDS-PAGE) as a single band with a molecular mass of 75kD.The native molecular mass determined by gel-filtration chromatography was 74 kDa, indicating that DELTA^1-tetrahydrocannabinolic acid synthase is a monomeric enzyme. Using isoelectric focusing, we determined a pI of 6.4 for the enzyme. The enzyme activity was greatest between pH5.5 and 6.0. The enzyme catalyzed the conversion of cannabigerolic acid or cannabinerolic acid to DELTA^1-tetrahydrocannabinolic acid. The V_<max> values were 2.68 and 0.37 nkat/mg for cannabigerolic acid and cannabinerolic acid, respectively. These results indicated that DELTA^1-tetrahydrocannabinolic acid is predominantly formed from cannabigerolic acid.

DELTA^1-四氢大麻酸合酶的存在在大麻（Cannabis sativa L.）的新鲜叶芽中得到证实。用大麻二酚酸测定的酶通过二乙氨基乙基纤维素、苯基琼脂糖CL-4B和羟基磷灰石色谱法纯化约300倍。纯化后的蛋白在十二烷基硫酸钠-聚丙烯酰胺凝胶电泳（SDS-PAGE）中迁移为单条带，分子量为75kD。凝胶过滤层析测定的天然分子量为74kDa，表明DELTA^1-四氢大麻酸合酶是单体酶。使用等电聚焦，我们确定该酶的等电点为 6.4。在pH5.5至6.0之间酶活性最大。该酶催化大麻二酚酸或大麻二酚酸转化为DELTA^1-四氢大麻酚酸。大麻二酚酸和大麻二酚酸的V_<max>值分别为2.68和0.37nkat/mg。这些结果表明DELTA^1-四氢大麻酚酸主要由大麻二酚酸形成。

项目成果

Futoshi Taura et al.: "New cannabinoid, cannabinerolic acid" from Cannabis sativa Phytochemistry. (in press). (1995)

Futoshi Taura 等人：“新大麻素，大麻酚酸”来自大麻植物化学。

FutoSHI TAURA: "A New Cannabinoids,Cannabinerlic Acid,from Cannabis Sativa" Phytochemistry. (1995)

FutoSHI TAURA：“一种新的大麻素，大麻二酸，来自大麻”植物化学。

FutoSHI TAURA: "A New Cannabinoid,Cannabinerolic acid,from Cannabis Sativa" Phytochemistry. (1995)

FutoSHI TAURA：“一种新的大麻素，来自大麻的大麻酚酸”植物化学。