Zheng Baohui, Zhu Shunying, Wu Xu
Cutaneous Biology Research Center, Massachusetts General Hospital, Harvard Medical School , Building 149, 13th Street, Charlestown, Massachusetts 02129, United States.
ACS Chem Biol. 2015 Jan 16;10(1):115-21. doi: 10.1021/cb500758s. Epub 2014 Oct 23.
Dynamic palmitoylation is an important post-translational modification regulating protein localization, trafficking, and signaling activities. The Asp-His-His-Cys (DHHC) domain containing enzymes are evolutionarily conserved palmitoyl acyltransferases (PATs) mediating diverse protein S-palmitoylation. Cerulenin is a natural product inhibitor of fatty acid biosynthesis and protein palmitoylation, through irreversible alkylation of the cysteine residues in the enzymes. Here, we report the synthesis and characterization of a "clickable" and long alkyl chain analogue of cerulenin as a chemical probe to investigate its cellular targets and to label and profile PATs in vitro and in live cells. Our results showed that the probe could stably label the DHHC-family PATs and enable mass spectrometry studies of PATs and other target proteins in the cellular proteome. Such probe provides a new chemical tool to dissect the functions of palmitoylating enzymes in cell signaling and diseases and reveals new cellular targets of the natural product cerulenin.
动态棕榈酰化是一种重要的翻译后修饰，可调节蛋白质的定位、运输和信号传导活性。含天冬氨酸-组氨酸-组氨酸-半胱氨酸（DHHC）结构域的酶是进化上保守的棕榈酰酰基转移酶（PATs），介导多种蛋白质的S-棕榈酰化。浅蓝菌素是一种脂肪酸生物合成和蛋白质棕榈酰化的天然产物抑制剂，它通过不可逆地烷基化酶中的半胱氨酸残基来发挥作用。在此，我们报告了一种浅蓝菌素的“可点击”且具有长烷基链的类似物的合成与表征，该类似物作为一种化学探针，用于研究其细胞靶点，并在体外和活细胞中标记和分析PATs。我们的结果表明，该探针能够稳定地标记DHHC家族的PATs，并能够对细胞蛋白质组中的PATs和其他靶蛋白进行质谱研究。这种探针为剖析棕榈酰化酶在细胞信号传导和疾病中的功能提供了一种新的化学工具，并揭示了天然产物浅蓝菌素的新细胞靶点。
