Enzymatic assay for perfluoro-tagged metabolites of l-DOPA using crude lysate from E. coli transformed with pKKAADCII.
Enzymatic assay for perfluoro-tagged metabolites of l-DOPA using crude lysate from E. coli transformed with pKKAADCII.
复制标题
使用 pKKAADCII 转化的大肠杆菌的粗裂解物对 L-DOPA 的全氟标记代谢物进行酶法测定。
DOI：
10.1007/s00253-003-1485-2
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发表时间：
2004
期刊：
影响因子：
--
通讯作者：
Guo,C
中科院分区：
文献类型：
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作者：
Dingman,S;Snyder-Leiby,T;Mack,DJ;Thomas,R;Guo,C
用质粒pKKAADC Ⅱ转化的大肠杆菌JM 109裂解液中的芳香酸脱羧酶（AADC）处理含有9个19 F原子标记的1-DOPA和多巴胺异构体。用HPLC分析所得样品。第一项研究使用标记的多巴胺作为标准，研究了标记的I-DOPA转化为标记的多巴胺。进行第二项研究，以确定在酶测定中看到的峰的来源。具有在位置5处结合的标签的I-DOPA充当AADC的最佳底物。适合AADC活性位点的异构体可能遵循多巴胺在体内的生物合成路径，并且在磁共振研究中可能有用。本文所述的酶测定法为筛选用于神经通路氟成像的新化合物提供了一种有效且具有成本效益的工具。
Isomers ofl-DOPA and dopamine with a nine-atom19F atom tag were exposed to aromatic acid decarboxylase (AADC) in the lysate ofEscherichia coliJM109 that had been transformed with the plasmid pKKAADCII. The resulting samples were analyzed with HPLC. The first study investigated the conversion of the taggedl-DOPA into tagged dopamine, using the tagged dopamine as a standard. A second study was undertaken to identify the source of peaks seen in the enzymatic assays.l-DOPA with the tag bonded at position 5 served as the best substrate for AADC. Isomers that fit into the active site of AADC are likely to follow the biosynthetic path for dopamine in vivo and are potentially useful in magnetic resonance studies. The enzymatic assay described here provides an efficient and cost-effective tool for screening new compounds for use in the fluorine imaging of neural pathways.