Abstract：
The (S)-norcoclaurine synthase from Thalictrum flavum (TfNCS) stereoselectively catalyzes the Pictet-Spengler reaction between dopamine and 4-hydroxyphenylacetaldehyde to give (S)-norcoclaurine. TfNCS can catalyze the Pictet-Spengler reaction with various aldehydes and ketones, leading to diverse tetrahydroisoquinolines. This substrate promiscuity positions TfNCS as a highly promising enzyme for synthesizing fine chemicals. Understanding carbonyl-containing substrates\' structural and electronic signatures that influence TfNCS activity can help expand its applications in the synthesis of different compounds and aid in protein optimization strategies. In this study, we investigated the influence of the molecular properties of aldehydes and ketones on their reactivity in the TfNCS-catalyzed Pictet-Spengler reaction. Initially, we compiled a library of reactive and unreactive compounds from previous publications. We also performed enzymatic assays using nuclear magnetic resonance to identify some reactive and unreactive carbonyl compounds, which were then included in the library. Subsequently, we employed QSAR and DFT calculations to establish correlations between substrate-candidate structures and reactivity. Our findings highlight correlations of structural and stereoelectronic features, including the electrophilicity of the carbonyl group, to the reactivity of aldehydes and ketones toward the TfNCS-catalyzed Pictet-Spengler reaction. Interestingly, experimental data of seven compounds out of fifty-three did not correlate with the electrophilicity of the carbonyl group. For these seven compounds, we identified unfavorable interactions between them and the TfNCS. Our results demonstrate the applications of in silico techniques in understanding enzyme promiscuity and specificity, with a particular emphasis on machine learning methodologies, DFT electronic structure calculations, and molecular dynamic (MD) simulations.
摘要：
来自黄thalictrum（TfNCS）的（S）-去甲氯胺合酶立体选择性催化多巴胺与4-羟基苯乙醛之间的Pictet-Spengler反应，得到（S）-去甲氯胺。TfNCS可以催化Pictet-Spengler与各种醛和酮的反应，导致不同的四氢异喹啉。这种底物混杂性使TfNCS成为合成精细化学品的非常有前途的酶。了解影响TfNCS活性的含羰基底物的结构和电子签名可以帮助扩展其在不同化合物合成中的应用，并有助于蛋白质优化策略。在这项研究中,在TfNCS催化的Pictet-Spengler反应中，我们研究了醛和酮的分子性质对其反应性的影响。最初，我们从以前的出版物中编译了一个反应性和非反应性化合物库。我们还使用核磁共振进行了酶测定，以鉴定一些反应性和非反应性羰基化合物，然后将其包含在图书馆中。随后，我们使用QSAR和DFT计算来建立底物候选结构和反应性之间的相关性。我们的发现强调了结构和立体电子特征的相关性，包括羰基的亲电性，醛和酮对TfNCS催化的Pictet-Spengler反应的反应性。有趣的是,53个化合物中的7个化合物的实验数据与羰基的亲电性无关。对于这七个化合物，我们确定了它们与TfNCS之间的不利相互作用。我们的结果证明了计算机技术在理解酶混杂性和特异性方面的应用，特别强调机器学习方法，DFT电子结构计算,和分子动力学(MD)模拟。
