Grothe T, Lenz R, Kutchan T M
Leibniz-Institut für Pflanzenbiochemie, Weinberg 3, 06120 Halle/Saale, Germany.
J Biol Chem. 2001 Aug 17;276(33):30717-23. doi: 10.1074/jbc.M102688200. Epub 2001 Jun 12.
Salutaridinol 7-O-acetyltransferase (EC ) catalyzes the conversion of the phenanthrene alkaloid salutaridinol to salutaridinol-7-O-acetate, the immediate precursor of thebaine along the morphine biosynthetic pathway. We have isolated a cDNA clone that corresponds to the internal amino acid sequences of the native enzyme purified from a cell suspension culture of opium poppy Papaver somniferum. The recombinant enzyme acetylated the 7-hydroxyl moiety of salutaridinol in the presence of acetyl-CoA. The apparent K(m) value for salutaridinol was determined to be 9 microm and 54 microm for acetyl-CoA. The gene transcript was detected in extracts from Papaver orientale and Papaver bracteatum in addition to P. somniferum. Genomic DNA gel blot analysis indicated that there is likely a single copy of this gene in the P. somniferum genome. The amino acid sequence of salutaridinol 7-O-acetyltransferase is most similar (37% identity) to that of deacetylvindoline acetyltransferase of Catharanthus roseus. Salutaridinol 7-O-acetyltransferase is the second enzyme specific to morphine biosynthesis for which we have isolated a cDNA. Taken together with the other cDNAs cloned encoding norcoclaurine 6-O-methyltransferase, (S)-N-methylcoclaurine 3'-hydroxylase, the cytochrome P-450 reductase, and codeinone reductase, significant progress has been made toward accumulating genes of this pathway to enable the end goal of a biotechnological production of morphinan alkaloids.
水杨醇苷醇7 - O - 乙酰基转移酶（EC ）催化菲类生物碱水杨醇苷醇转化为水杨醇苷醇 - 7 - O - 乙酸酯，后者是吗啡生物合成途径中蒂巴因的直接前体。我们从罂粟（Papaver somniferum）的细胞悬浮培养物中纯化了天然酶，并根据其内部氨基酸序列分离出了一个cDNA克隆。重组酶在乙酰辅酶A存在的情况下，将水杨醇苷醇的7 - 羟基部分乙酰化。水杨醇苷醇的表观K(m)值测定为9微摩尔，乙酰辅酶A的表观K(m)值为54微摩尔。除了罂粟外，在东方罂粟（Papaver orientale）和苞叶罂粟（Papaver bracteatum）的提取物中也检测到了该基因转录本。基因组DNA凝胶印迹分析表明，在罂粟基因组中该基因可能只有一个拷贝。水杨醇苷醇7 - O - 乙酰基转移酶的氨基酸序列与长春花（Catharanthus roseus）的去乙酰文朵灵乙酰转移酶最为相似（同一性为37%）。水杨醇苷醇7 - O - 乙酰基转移酶是我们分离出cDNA的第二种吗啡生物合成特异性酶。连同其他已克隆的编码去甲乌药碱6 - O - 甲基转移酶、（S）- N - 甲基乌药碱3'- 羟化酶、细胞色素P - 450还原酶和可待因酮还原酶的cDNA，在积累该途径的基因以实现吗啡烷生物碱生物技术生产的最终目标方面已经取得了重大进展。
