Chemistry Research Laboratory, Department of Chemistry, University of Oxford, Oxford, UK.
Department of Chemistry, Wayne State University, Detroit, MI, USA.
Nature. 2020 Sep;585(7826):530-537. doi: 10.1038/s41586-020-2733-7. Epub 2020 Sep 23.
Post-translational modifications (PTMs) greatly expand the structures and functions of proteins in nature. Although synthetic protein functionalization strategies allow mimicry of PTMs, as well as formation of unnatural protein variants with diverse potential functions, including drug carrying, tracking, imaging and partner crosslinking, the range of functional groups that can be introduced remains limited. Here we describe the visible-light-driven installation of side chains at dehydroalanine residues in proteins through the formation of carbon-centred radicals that allow C-C bond formation in water. Control of the reaction redox allows site-selective modification with good conversions and reduced protein damage. In situ generation of boronic acid catechol ester derivatives generates RHC radicals that form the native (β-CH-γ-CH) linkage of natural residues and PTMs, whereas in situ potentiation of pyridylsulfonyl derivatives by Fe(II) generates RFC radicals that form equivalent β-CH-γ-CF linkages bearing difluoromethylene labels. These reactions are chemically tolerant and incorporate a wide range of functionalities (more than 50 unique residues/side chains) into diverse protein scaffolds and sites. Initiation can be applied chemoselectively in the presence of sensitive groups in the radical precursors, enabling installation of previously incompatible side chains. The resulting protein function and reactivity are used to install radical precursors for homolytic on-protein radical generation; to study enzyme function with natural, unnatural and CF-labelled post-translationally modified protein substrates via simultaneous sensing of both chemo- and stereoselectivity; and to create generalized 'alkylator proteins' with a spectrum of heterolytic covalent-bond-forming activity (that is, reacting diversely with small molecules at one extreme or selectively with protein targets through good mimicry at the other). Post-translational access to such reactions and chemical groups on proteins could be useful in both revealing and creating protein function.
翻译后修饰（PTMs）极大地扩展了自然界中蛋白质的结构和功能。尽管合成蛋白质功能化策略能够模拟PTMs，并形成具有多种潜在功能的非天然蛋白质变体，包括药物携带、追踪、成像和伴侣交联，但可引入的官能团范围仍然有限。在此，我们描述了通过形成以碳为中心的自由基，在蛋白质的脱氢丙氨酸残基上可见光驱动安装侧链，该自由基能够在水中形成碳-碳键。控制反应的氧化还原过程可实现位点选择性修饰，具有良好的转化率并减少蛋白质损伤。原位生成硼酸儿茶酚酯衍生物会产生RHC自由基，形成天然残基和PTMs的天然（β-CH-γ-CH）连接，而Fe(II)对吡啶基磺酰基衍生物的原位增强作用会产生RFC自由基，形成带有二氟亚甲基标签的等效β-CH-γ-CF连接。这些反应具有化学耐受性，可将多种功能基团（超过50种独特的残基/侧链）引入不同的蛋白质支架和位点。引发反应可以在自由基前体中存在敏感基团的情况下进行化学选择性应用，从而能够安装先前不相容的侧链。利用所得蛋白质的功能和反应活性来安装用于均裂蛋白质自由基生成的自由基前体；通过同时感知化学选择性和立体选择性，使用天然、非天然和CF标记的翻译后修饰蛋白质底物来研究酶的功能；并创建具有一系列异裂共价键形成活性的广义“烷基化蛋白质”（即在一个极端与小分子进行多样化反应，或在另一个极端通过良好的模拟与蛋白质靶点进行选择性反应）。对蛋白质上此类反应和化学基团的翻译后访问在揭示和创造蛋白质功能方面都可能有用。
