Rosmarinic acid synthase is a new member of the superfamily of BAHD acyltransferases
Rosmarinic acid synthase is a new member of the superfamily of BAHD acyltransferases
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DOI：
10.1007/s00425-006-0393-y
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发表时间：
2006-11-01
期刊：
影响因子：
4.3
通讯作者：
Petersen, Maike
中科院分区：
文献类型：
--
作者：
Berger, Anja;Meinhard, Juliane;Petersen, Maike
彩叶草悬浮细胞中迷迭香酸合成酶的纯化（唇形科）通过分级硫酸铵沉淀、疏水相互作用层析和两个亲和层析步骤鉴定了肽序列，这使得基于PCR的方法能够分离编码该酶的全长cDNA。该cDNA的开放阅读框长度为1290个碱基对，编码430个氨基酸残基的蛋白质，分子量为47，932 Da，具有BAHD超家族酰基转移酶的典型特征。该cDNA在大肠杆菌中异源表达。该酶使用4-香豆酰-和咖啡酰-辅酶A和4-羟基苯乳酸以及3.4-二羟基苯乳酸作为底物显示迷迭香酸合成酶的活性。莽草酸和奎尼酸不能作为羟基肉桂酰受体。因此，这是迷迭香酸合成酶的cDNA克隆的第一个报告。
Purification of rosmarinic acid synthase (hydroxycinnamoyl-CoA:hydroxyphenyllactate hydroxycinnamoyltransferase) from suspension cells of Coleus blumei Benth. (Lamiaceae) by fractionated ammonium sulphate precipitation, hydrophobic interaction chromatography and two affinity chromatography steps led to the identification of peptide sequences, which enabled a PCR-based approach to isolate the full-length cDNA encoding this enzyme. The open reading frame of the cDNA had a length of 1290 base pairs encoding a protein of 430 amino acid residues with a molecular mass of 47,932 Da with typical characteristics of an acyltransferase of the BAHD superfamily. The cDNA was heterologously expressed in Escherichia coli. The enzyme displayed the activity of rosmarinic acid synthase using 4-coumaroyl- and caffeoyl-coenzyme A and 4-hydroxyphenyllactate as well as 3.4-dihydroxyphenyllactate as substrates. Shikimic acid and quinic acid were not able to serve as hydroxycinnamoyl acceptors. This therefore is the first report of the cDNA-cloning of a rosmarinic acid synthase.