Natural metalloproteins perform many functions — ranging from sensing to electron transfer and catalysis — in which the position and property of each ligand and metal are dictated by protein structure. De novo protein design aims to define an amino acid sequence that encodes a specific structure and function, providing a critical test of the hypothetical inner workings of (metallo)proteins. To date, de novo metalloproteins have used simple, symmetric tertiary structures — uncomplicated by the large size and evolutionary marks of natural proteins — to interrogate structure–function hypotheses. In this Review, we discuss de novo design applications, such as proteins that induce complex, increasingly asymmetric ligand geometries to achieve function, as well as the use of more canonical ligand geometries to achieve stability. De novo design has been used to explore how proteins fine-tune redox potentials and catalyse both oxidative and hydrolytic reactions. With an increased understanding of structure–function relationships, functional proteins including O₂-dependent oxidases, fast hydrolases and multi-proton/multielectron reductases have been created. In addition, proteins can now be designed using xenobiological metals or cofactors and principles from inorganic chemistry to derive new-to-nature functions. These results and the advances in computational protein design suggest a bright future for the de novo design of diverse, functional metalloproteins.

天然金属蛋白具有多种功能——从传感到电子转移和催化——其中每个配体和金属的位置和性质由蛋白质结构决定。从头蛋白质设计旨在定义编码特定结构和功能的氨基酸序列，为（金属）蛋白质的假设内部运作提供关键测试。迄今为止，从头金属蛋白已使用简单、对称的三级结构（天然蛋白质的大尺寸和进化标记并不复杂）来质疑结构功能假设。在这篇综述中，我们讨论了从头设计应用，例如诱导复杂、越来越不对称的配体几何形状以实现功能的蛋白质，以及使用更规范的配体几何形状来实现稳定性。从头设计已用于探索蛋白质如何微调氧化还原电位并催化氧化和水解反应。随着对结构与功能关系的了解不断加深，包括 O ₂依赖性氧化酶、快速水解酶和多质子/多电子还原酶在内的功能蛋白已被创建。此外，现在可以使用异种生物金属或辅因子以及无机化学原理来设计蛋白质，以获得新的自然功能。这些结果和计算蛋白质设计的进展表明，多样化功能金属蛋白的从头设计有着光明的前景。