L-DOPA Dioxygenase Activity on 6-Substituted Dopamine Analogues
L-DOPA Dioxygenase Activity on 6-Substituted Dopamine Analogues
复制标题
DOI：
10.1021/acs.biochem.1c00310
复制
发表时间：
2021-07-29
期刊：
影响因子：
2.9
通讯作者：
Peterson, Larryn W.
中科院分区：
文献类型：
--
作者：
Goldberg, Alexander M.;Robinson, Miranda K.;Peterson, Larryn W.
双加氧酶是分解植物木本组织结构成分儿茶酚环的重要蛋白质催化剂。这种强大的化学作用在自然界中被用于制造抗生素和其他生物活性材料或降解植物材料，但我们对这些强大催化剂的底物空间的广度和深度的了解有限。本文报道了在6位取代的多巴胺衍生物作为左旋多巴双加氧酶底物的稳态和预稳态动力学分析，并分析了取代基吸电子性质的功能。稳态和预稳态动力学数据表明，多巴胺在与共底物分子氧的结合和催化作用中受损，这可能提供了早期反应中间体多巴胺半醌的光谱观察。多巴胺在前稳定状态下的反应途径与活性部位氧结合的非生产性模式一致。尽管存在这些限制，但左旋多巴双加氧酶能够结合所有多巴胺衍生物，并催化多巴胺、6-溴多巴胺、6-羧基多巴胺和6-氰多巴胺的多次环裂解。6-硝基多巴胺是单次转化的底物。该酶接受的底物种类与多种因素的相互作用一致，包括活性位点结合6位大负电荷基团的能力和每种儿茶酚胺的整体氧化性，这表明了外二醇裂解在半合成和生物修复中的应用。
Dioxygenase enzymes are essential protein catalysts for the breakdown of catecholic rings, structural components of plant woody tissue. This powerful chemistry is used in nature to make antibiotics and other bioactive materials or degrade plant material, but we have a limited understanding of the breadth and depth of substrate space for these potent catalysts. Here we report steady-state and pre-steady-state kinetic analysis of dopamine derivatives substituted at the 6-position as substrates of L-DOPA dioxygenase, and an analysis of that activity as a function of the electron-withdrawing nature of the substituent. Steady-state and pre-steady-state kinetic data demonstrate the dopamines are impaired in binding and catalysis with respect to the cosubstrate molecular oxygen, which likely afforded spectroscopic observation of an early reaction intermediate, the semiquinone of dopamine. The reaction pathway of dopamine in the pre-steady state is consistent with a nonproductive mode of binding of oxygen at the active site. Despite these limitations, L-DOPA dioxygenase is capable of binding all of the dopamine derivatives and catalyzing multiple turnovers of ring cleavage for dopamine, 6-bromodopamine, 6-carboxydopamine, and 6-cyanodopamine. 6-Nitrodopamine was a single-turnover substrate. The variety of substrates accepted by the enzyme is consistent with an interplay of factors, including the capacity of the active site to bind large, negatively charged groups at the 6-position and the overall oxidizability of each catecholamine, and is indicative of the utility of extradiol cleavage in semisynthetic and bioremediation applications.