Abstract

Deoxyribose-5-phosphate aldolases (DERAs, EC 4.1.2.4) are acetaldehyde-dependent, Class I aldolases catalyzing in nature a reversible aldol reaction between an acetaldehyde donor (C2 compound) and glyceraldehyde-3-phosphate acceptor (C3 compound, C3P) to generate deoxyribose-5-phosphate (C5 compound, DR5P). DERA enzymes have been found to accept also other types of aldehydes as their donor, and in particular as acceptor molecules. Consequently, DERA enzymes can be applied in C–C bond formation reactions to produce novel compounds, thus offering a versatile biocatalytic alternative for synthesis. DERA enzymes, found in all kingdoms of life, share a common TIM barrel fold despite the low overall sequence identity. The catalytic mechanism is well-studied and involves formation of a covalent enzyme-substrate intermediate. A number of protein engineering studies to optimize substrate specificity, enzyme efficiency, and stability of DERA aldolases have been published. These have employed various engineering strategies including structure-based design, directed evolution, and recently also machine learning–guided protein engineering. For application purposes, enzyme immobilization and usage of whole cell catalysis are preferred methods as they improve the overall performance of the biocatalytic processes, including often also the stability of the enzyme. Besides single-step enzymatic reactions, DERA aldolases have also been applied in multi-enzyme cascade reactions both in vitro and in vivo. The DERA-based applications range from synthesis of commodity chemicals and flavours to more complicated and high-value pharmaceutical compounds.

Key points

• DERA aldolases are versatile biocatalysts able to make new C–C bonds.

• Synthetic utility of DERAs has been improved by protein engineering approaches.

• Computational methods are expected to speed up the future DERA engineering efforts.

Graphical abstract

中文翻译：

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2-脱氧-D-核糖5-磷酸醛缩酶酶工程的现状和需求及其影响

摘要

5-磷酸脱氧核糖醛缩酶（DERA，EC 4.1.2.4）是乙醛依赖性的 I 类醛缩酶，在自然界中催化乙醛供体（C2 化合物）和 3-磷酸甘油醛受体（C3 化合物，C3P）之间的可逆醛缩醛反应生成脱氧核糖 5-磷酸（C5 化合物，DR5P）。已经发现DERA酶也接受其他类型的醛作为它们的供体，特别是作为受体分子。因此，DERA 酶可用于 C-C 键形成反应以产生新化合物，从而为合成提供通用的生物催化替代方案。尽管总体序列同一性较低，但在所有生命王国中都发现的 DERA 酶具有共同的 TIM 桶折叠。催化机制已得到充分研究，涉及共价酶-底物中间体的形成。已经发表了许多优化 DERA 醛缩酶的底物特异性、酶效率和稳定性的蛋白质工程研究。它们采用了各种工程策略，包括基于结构的设计、定向进化，以及最近机器学习引导的蛋白质工程。出于应用目的，酶固定化和全细胞催化的使用是优选的方法，因为它们改善了生物催化过程的整体性能，通常还包括酶的稳定性。除了单步酶促反应外，DERA 醛缩酶还被应用于体外和体内的多酶级联反应。基于DERA的应用范围从商品化学品和香料的合成到更复杂和高价值的药物化合物。

关键点

• DERA 醛缩酶是一种通用的生物催化剂，能够形成新的 C-C 键。

• 蛋白质工程方法改善了DERA 的合成效用。

• 计算方法有望加速未来的DERA 工程工作。

图形概要