Identification of the dioxygenase-generated intermediate formed during biosynthesis of the dihydropyrrole moiety common to anthramycin and sibiromycin.
Identification of the dioxygenase-generated intermediate formed during biosynthesis of the dihydropyrrole moiety common to anthramycin and sibiromycin.
复制标题
鉴定在蒽霉素和西伯霉素共有的二氢吡咯部分生物合成过程中形成的双加氧酶产生的中间体。
DOI：
10.1016/j.bmc.2014.12.024
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发表时间：
2015
影响因子：
3.5
通讯作者：
Rokita,StevenE
中科院分区：
文献类型：
--
作者：
Saha,Shalini;Li,Wei;Gerratana,Barbara;Rokita,StevenE
吡咯并[1,4]苯二氮卓（PBD）生物合成的描述是工程生产具有增强抗肿瘤活性的类似物的先决条件。预测的双加氧酶 Orf12 和 SibV 分别与 PBD 蒽霉素和西伯霉素中的二氢吡咯生物合成相关，并进行表达和纯化用于活性研究。紫外可见光谱显示这些酶催化 1-3,4-二羟基苯丙氨酸 (l-DOPA) 的区域特异性 2,3-extradiol 双氧化反应形成 1-2,3-secodopa (λmax= 368 nm)。 1H NMR 光谱表明 1-2,3-secodopa 环化为 α-酮酸互变异构体1-4-(2-氧代-3-丁烯酸)-4,5-二氢吡咯-2-羧酸 (λmax= 414 nm)。因此，双加氧酶是建立二氢吡咯部分支架的关键。动力学研究表明双加氧酶产物相对不稳定，可能会被后续的生物合成步骤迅速消耗。这些双加氧酶的酶产物和二聚体状态在PBD和非PBD途径中参与二氢吡咯和吡咯烷生物合成的双加氧酶中是保守的。
A description of pyrrolo[1,4]benzodiazepine (PBD) biosynthesis is a prerequisite for engineering production of analogs with enhanced antitumor activity. Predicted dioxygenases Orf12 and SibV associated with dihydropyrrole biosynthesis in PBDs anthramycin and sibiromycin, respectively, were expressed and purified for activity studies. UV–visible spectroscopy revealed that these enzymes catalyze the regiospecific 2,3-extradiol dioxygenation ofl-3,4-dihydroxyphenylalanine (l-DOPA) to forml-2,3-secodopa (λmax= 368 nm).1H NMR spectroscopy indicates thatl-2,3-secodopa cyclizes into the α-keto acid tautomer ofl-4-(2-oxo-3-butenoic-acid)-4,5-dihydropyrrole-2-carboxylic acid (λmax= 414 nm). Thus, the dioxygenases are key for establishing the scaffold of the dihydropyrrole moiety. Kinetic studies suggest the dioxygenase product is relatively labile and is likely consumed rapidly by subsequent biosynthetic steps. The enzymatic product and dimeric state of these dioxygenases are conserved in dioxygenases involved in dihydropyrrole and pyrrolidine biosynthesis within both PBD and non-PBD pathways.