Applied Biotechnology Group, European University of Madrid, Villaviciosa de Odón, Spain.
Institute of Theoretical Chemistry, Faculty of Chemistry, Währinger Str. 17, A-1090 University of Vienna, Vienna, Austria.
Org Biomol Chem. 2019 Aug 28;17(34):7891-7899. doi: 10.1039/c9ob01315f.
Insight into the catalytic mechanism of Lactobacillus leichmannii nucleoside 2'-deoxyribosyltransferase (LlNDT) has been gained by calculating a quantum mechanics-molecular mechanics (QM/MM) free-energy landscape of the reaction within the enzyme active site. Our results support an oxocarbenium species as the reaction intermediate and thus an SN1 reaction mechanism in this family of bacterial enzymes. Our mechanistic proposal is validated by comparing experimental kinetic data on the impact of the single amino acid replacements Tyr7, Glu98 and Met125 with Ala, Asp and Ala/norLeu, respectively, and accounts for the specificity shown by this enzyme on a non-natural substrate. This work broadens our understanding of enzymatic C-N bond cleavage and C-N bond formation.
通过在酶活性位点内计算反应的量子力学-分子力学（QM/MM）自由能景观，深入了解了乳酸乳球菌核苷 2'-脱氧核糖基转移酶（LlNDT）的催化机制。我们的结果支持氧碳正离子物种作为反应中间体，因此在该家族的细菌酶中为 SN1 反应机制。我们的机理假设通过比较实验动力学数据得到了验证，该数据涉及对单个氨基酸替换 Tyr7、Glu98 和 Met125 分别用 Ala、Asp 和 Ala/norLeu 的影响，并且解释了该酶在非天然底物上表现出的特异性。这项工作拓宽了我们对酶 C-N 键断裂和 C-N 键形成的理解。
