The Francis Crick Institute, London, UK.
Imperial College London, Department of Chemistry, Molecular Sciences Research Hub, London, UK.
Nat Biotechnol. 2024 Oct;42(10):1548-1558. doi: 10.1038/s41587-023-02030-0. Epub 2024 Jan 8.
The 23 human zinc finger Asp-His-His-Cys motif-containing (ZDHHC) S-acyltransferases catalyze long-chain S-acylation at cysteine residues across an extensive network of hundreds of proteins important for normal physiology or dysregulated in disease. Here we present a technology to directly map the protein substrates of a specific ZDHHC at the whole-proteome level, in intact cells. Structure-guided engineering of paired ZDHHC 'hole' mutants and 'bumped' chemically tagged fatty acid probes enabled probe transfer to specific protein substrates with excellent selectivity over wild-type ZDHHCs. Chemical-genetic systems were exemplified for five human ZDHHCs (3, 7, 11, 15 and 20) and applied to generate de novo ZDHHC substrate profiles, identifying >300 substrates and S-acylation sites for new functionally diverse proteins across multiple cell lines. We expect that this platform will elucidate S-acylation biology for a wide range of models and organisms.
23 个人类锌指 Asp-His-His-Cys 基序含有（ZDHHC）S-酰基转移酶催化长链 S-酰化在数百个对正常生理很重要的蛋白质的半胱氨酸残基上，或在疾病中失调。在这里，我们提出了一种在完整细胞中直接在全蛋白质组水平上绘制特定 ZDHHC 的蛋白质底物的技术。结构引导的配对 ZDHHC“孔”突变体和“凸起”化学标记脂肪酸探针的工程化，使探针能够与特定的蛋白质底物转移，对野生型 ZDHHC 具有极好的选择性。化学遗传系统的例子有五个人类 ZDHHC（3、7、11、15 和 20），并应用于生成新的功能多样化蛋白质的从头 ZDHHC 底物谱，鉴定了多个细胞系中的 >300 个底物和 S-酰化位点。我们预计，该平台将阐明广泛模型和生物体的 S-酰化生物学。
