Halogenases in Biocatalysis: Advances in Mechanism Elucidation， Directed Evolution， and Green Manufacturing Synthetic Biology Journal    DOI: 10.12211/2096-8280.2024-091

Fig. 7 The typical structure of single-component flavin-dependent halogenases

Extracts from the Article

然而，外源黄素还原酶的需求导致FDHs催化体系复杂化，显著增加了反应成本。值得关注的是，自然界微生物资源中存在单组份卤化酶（Single-component FDH，ScFDH），这类酶通过单一蛋白整合还原酶与卤化酶的双重功能，可自主完成黄素再生与卤素转移的偶联反应。目前，发现两种ScFDH，一种是从海洋细菌Pseudoalteromonas leteoviolacea中分离得到的Bmp5［65］，另一种是从水生细菌Aetokthonos hydrillicola中分离的AetF［66］。这两种酶都是单体，与之前研究的FDH存在显著的结构差异。与常见的方盒-金字塔结构不同，Bmp5和AetF的结构轮廓类似于澳洲大陆［67-68］（图7）。如图7所示，Bmp5和AetF底部存在被随机卷曲基序环绕的较大空腔，允许NADPH和FADH₂进入并与酶结合。Bmp5和AetF的轮廓及内部功能区域高度重叠，包括活性位点（Bmp5-K277，AetF-K258），显示了ScFDH的一般结构特征。最近，Tenebro等［38］报道了ScFDH家族的新成员ChlCz9（UNT00659.1），分离自链霉菌Streptomyces tubbatahanensis。然而，根据氨基酸序列比对和AlphaFold建模，笔者发现ChlCz9的预测结构与Bmp5和AetF显著不同，但与PltM高度相似（图7）。序列比对结果也显示ChlCz9缺乏结合NADP的基本元件。因此，需要进一步通过直接活性测定或高分辨率晶体结构分析，确认ChlCz9是否为新型ScFDH。

Other Images/Table from this Article

• Fig. 1  Halogens are the most common and important modification groups in pesticides and pharmaceutical products
• Fig. 2  Examples of halogenated natural products and the role of halogenases in molecular synthesis^[4,12,15-16]
• Fig. 3  Cofactors and catalytic mechanisms of different types of halogenase^[1,4,17-25]
• Fig. 4  Workflow of genome mining for halogenases and their application examples^[34]
• Table 1  Examples of halogenase discovered by genome mining
• Table 2  The uniqueness and superiority of the CLEAN algorithm framework
• Fig. 5  Overview of chloride depletion workflow for revealing cryptic halogenation^[59]
• Fig. 6  The typical structure and catalytic elements of flavin-dependent halogenases
• Fig. 8  The proposed catalytic mechanism of PrnA by calculation and simulation^[69]
• Fig. 9  The typical structure of NHFe/αKGHs^[25,70-76]
• Fig. 10  The proposed catalytic mechanism of BesD by calculation and simulation^[80]
• Fig. 11  The action mode and engineering strategies of novel nucleotide halogenase ^[46]
• Fig.12  The structural, function and catalytic mechanism of copper-dependent halogenase ApnU^[81]
• Table 3  Typical engineering cases of natural halogenase in recent years
• Fig. 13  The typical structure SAM-dependent halogenase ^[84-91]The red lines indicate the residues that are crucial for fluorination activity; the yellow lines indicate the residues that can form hydrogen bonds with the modification groups at the C2′ position of adenine
• Fig. 14  Tunnel engineering improves the catalytic performance of Thal halogenase^[96]
• Fig. 15  Directed evolution enhances the fluorination activity of non-heme iron oxidase^[108-109]
• Fig. 16  Biocatalytic synthesis platform of halogenated tryptophan derivatives^[113]
• Tab. 4  Examples of FDHs catalyzing enantioselective reactions^[125-127]
• Tab. 5  Recent cases and key data related to the potential industrial application of halogenases
• Fig. 17  Representative research progress in the field of halogenase research
• Fig. 18  AI/ML drives the intelligent evolution of halogenase^[138-156]