Chemical Proteomic Profiling of the Interacting Proteins of Isoprenoid Pyrophosphates.
Chemical Proteomic Profiling of the Interacting Proteins of Isoprenoid Pyrophosphates.
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DOI：
10.1021/acs.analchem.0c01676
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发表时间：
2020-05
影响因子：
7.4
通讯作者：
Rong Cai;Xuejiao Dong;Kailin Yu;Xiaomei He;Xiaochuan Liu;Yinsheng Wang
中科院分区：
文献类型：
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作者：
Rong Cai;Xuejiao Dong;Kailin Yu;Xiaomei He;Xiaochuan Liu;Yinsheng Wang
类异戊二烯焦磷酸参与蛋白质的异戊二烯基化，并在细胞中发挥调节作用；然而，对能与类异戊二烯焦磷酸相互作用的细胞蛋白知之甚少。在这里，我们设计了一种化学蛋白质组学策略，利用脱硫生物素-香叶基焦磷酸(GPP)酰基磷酸探针，用于浓缩和随后使用LC-MS/MS(LC-MS/MS)鉴定GPP结合蛋白。通过结合细胞培养中氨基酸的稳定同位素标记(SILAC)和低浓度与高浓度GPP探针的竞争标记，与ATP与GPP酰基磷酸探针的竞争标记，或者在不同浓度的游离GPP存在的情况下与GPP探针的竞争标记，我们发现了一些候选的GPP结合蛋白。我们还首次发现组蛋白脱乙酰基酶1(HDAC1)是一种GPP结合蛋白。此外，我们还发现类异戊二烯焦磷酸可以调节HDAC1的酶活性。我们共同开发了一种新的化学蛋白质组学方法，用于在蛋白质组范围内发现GPP结合蛋白，这为更好地理解异戊二烯类化合物的生物学功能奠定了基础。
Isoprenoid pyrophosphates are involved in protein prenylation and assume regulatory roles in cells; however, little is known about the cellular proteins that can interact with isoprenoid pyrophosphates. Here, we devised a chemical proteomic strategy, capitalizing on the use of desthiobiotin-geranyl pyrophosphate (GPP) acyl phosphate probe, for the enrichment and subsequent identification of GPP-binding proteins using liquid chromatography-tandem mass spectrometry (LC-MS/MS). By combining stable isotope labeling by amino acids in cell culture (SILAC) and competitive labeling with low vs. high concentrations of GPP probe, with ATP vs. GPP acyl phosphate probes, or with the GPP probe in the presence of different concentrations of free GPP, we uncovered a number of candidate GPP-binding proteins. We also discovered, for the first time, histone deacetylase 1 (HDAC1) as a GPP-binding protein. Furthermore, we found that the enzymatic activity of HDAC1 could be modulated by isoprenoid pyrophosphates. Together, we developed a novel chemical proteomic method for the proteome-wide discovery of GPP-binding proteins, which sets the stage for a better understanding about the biological functions of isoprenoids.