Zhao Luyao, Thongrakon Bhu-Bhud, Gautom Trishnamoni, Sahlberg Viktor, Berglund Per
Department of Industrial Biotechnology, School of Engineering Sciences in Chemistry, Biotechnology and Health, KTH Royal Institute of Technology, SE-10691, Stockholm, Sweden.
Chembiochem. 2025 Jul 11;26(13):e202500155. doi: 10.1002/cbic.202500155. Epub 2025 May 30.
Amine transaminases (ATAs), belonging to the class III transaminases within the superfamily of pyridoxal-5'-phosphate-dependent enzymes, catalyze transamination reactions between amino donors and amino acceptors. These enzymes are particularly appealing for their role in stereospecific synthesis of chiral amines. However, the stability of most ATAs is not satisfying, limiting their suitability for industrial applications. Among them, the amine transaminase from Silicibacter pomeroyi (Sp-ATA) has drawn attention due to its high activity and broad substrate scope under mild conditions and high pH. Nevertheless, maintaining the activity at higher temperatures is a challenge. Previous studies to enhance enzyme function through directed evolution have shown promising results, yet predicting the cooperative effects of individual stabilizing mutations remains challenging. An alternative strategy is ancestral sequence reconstruction (ASR), which is based on gene sequences to create a more or less artificial phylogenetic tree. This study aims to leverage ASR techniques to explore the thermostability, solvent tolerance, and substrate profile of Sp-ATA, to find more stable transaminases. By using Sp-ATA as a template and incorporating insights from ancestral sequences, this strategy offers a promising approach for developing robust biocatalysts suitable for industrial applications.
胺转氨酶（ATA）属于磷酸吡哆醛依赖性酶超家族中的III类转氨酶，催化氨基供体和氨基受体之间的转氨反应。这些酶因其在手性胺的立体特异性合成中的作用而特别具有吸引力。然而，大多数ATA的稳定性并不令人满意，限制了它们在工业应用中的适用性。其中，来自波氏硅杆菌的胺转氨酶（Sp-ATA）因其在温和条件和高pH下具有高活性和广泛的底物范围而受到关注。然而，在较高温度下保持活性是一项挑战。先前通过定向进化增强酶功能的研究已显示出有希望的结果，但预测单个稳定突变的协同效应仍然具有挑战性。另一种策略是祖先序列重建（ASR），它基于基因序列创建一个或多或少的人工系统发育树。本研究旨在利用ASR技术探索Sp-ATA的热稳定性、耐溶剂性和底物谱，以找到更稳定的转氨酶。通过以Sp-ATA为模板并结合祖先序列的见解，该策略为开发适用于工业应用的强大生物催化剂提供了一种有前途的方法。
