Warshel Institute for Computational Biology, School of Medicine, The Chinese University of Hong Kong, Shenzhen, 518172, Guangdong, China.
Kobilka Institute of Innovative Drug Discovery, School of Medicine, The Chinese University of Hong Kong, Shenzhen, 518172, Guangdong, China.
Nat Commun. 2024 Jun 19;15(1):5254. doi: 10.1038/s41467-024-49147-7.
C2'-halogenation has been recognized as an essential modification to enhance the drug-like properties of nucleotide analogs. The direct C2'-halogenation of the nucleotide 2'-deoxyadenosine-5'-monophosphate (dAMP) has recently been achieved using the Fe(II)/α-ketoglutarate-dependent nucleotide halogenase AdaV. However, the limited substrate scope of this enzyme hampers its broader applications. In this study, we report two halogenases capable of halogenating 2'-deoxyguanosine monophosphate (dGMP), thereby expanding the family of nucleotide halogenases. Computational studies reveal that nucleotide specificity is regulated by the binding pose of the phosphate group. Based on these findings, we successfully engineered the substrate specificity of these halogenases by mutating second-sphere residues. This work expands the toolbox of nucleotide halogenases and provides insights into the regulation mechanism of nucleotide specificity.
C2'-卤化已被认为是增强核苷酸类似物药物性质的重要修饰方式。最近，使用 Fe(II)/α-酮戊二酸依赖性核苷酸卤化酶 AdaV 实现了核苷酸 2'-脱氧腺苷-5'-单磷酸（dAMP）的直接 C2'-卤化。然而，该酶的有限底物范围限制了其更广泛的应用。在这项研究中，我们报告了两种能够卤化 2'-脱氧鸟苷单磷酸（dGMP）的卤化酶，从而扩展了核苷酸卤化酶家族。计算研究表明，核苷酸特异性受磷酸基团的结合构象调节。基于这些发现，我们通过突变第二壳层残基成功地对这些卤化酶的底物特异性进行了工程改造。这项工作扩展了核苷酸卤化酶的工具包，并提供了对核苷酸特异性调节机制的深入了解。
