Department of Chemistry, Gettysburg College, 300 N. Washington Street, Gettysburg, PA, 17325, USA.
Department of Chemistry, Ursinus College, 601 E Main Street, Collegeville, PA, 19426, USA.
J Biol Inorg Chem. 2021 Feb;26(1):161-167. doi: 10.1007/s00775-020-01845-5. Epub 2021 Jan 19.
Metallohydrolases are broadly used throughout biology, often to catalyze the degradation of macromolecules such as DNA and proteins. Many of these enzymes function with zinc in their active site, and an important subset of these enzymes utilize a binuclear zinc active site. Mimics of these enzymes have been developed, some of which catalyze the digestion of DNA. However, the majority of the mimics that utilize zinc are small molecules, and most are mononuclear. Herein, we report DNA cleavage activity by the de novo designed Due Ferri single-chain (DFsc) protein containing a binuclear zinc active site. This binuclear zinc-protein complex is able to digest plasmid DNA at rates up to 50 ng/h, and these cleavage rates are affected by changes to amino acid residues near the zinc-binding site. These results indicate that the DFsc scaffold is a good model system to carry out careful structure-function relationship studies to understand key structural features that influence reactivity in natural binuclear zinc hydrolases, as it is the first report of a binuclear model system in a protein scaffold.
金属水解酶在生物学中被广泛应用，常用于催化 DNA 和蛋白质等大分子的降解。这些酶中的许多在其活性部位都含有锌，而其中一个重要的亚类则利用双核锌活性部位。这些酶的模拟物已经被开发出来，其中一些可以催化 DNA 的消化。然而，利用锌的模拟物大多数是小分子，且大多数是单核的。本文报道了一种新设计的含有双核锌活性部位的双铁单链（DFsc）蛋白的 DNA 切割活性。这种双核锌-蛋白复合物能够以高达 50ng/h 的速度消化质粒 DNA，并且这些切割速率受到锌结合位点附近氨基酸残基变化的影响。这些结果表明，DFsc 支架是一个很好的模型系统，可以进行仔细的结构-功能关系研究，以了解影响自然双核锌水解酶反应性的关键结构特征，因为这是首次在蛋白质支架中报道双核模型系统。
