Department of Chemical Biology, Max-Planck-Institute of, Molecular Physiology, Otto-Hahn-Strasse 11, 44227, Dortmund, Germany.
Faculty of Chemistry and Chemical Biology, TU Dortmund University, Otto-Hahn-Strasse 6, 44227, Dortmund, Germany.
Chembiochem. 2019 Jan 18;20(2):134-139. doi: 10.1002/cbic.201800406. Epub 2018 Oct 17.
Protein myristoylation plays key roles in biological processes, for instance, in membrane attachment and activation of proteins and in mediating protein-protein and protein-lipid interactions. Furthermore, myristoylated proteins are involved in disorders, including cancer and viral infections. Therefore, new tools to study protein myristoylation are in high demand. Herein, we report the development of photoactivatable probes, based on a diazirine-substituted analogue of myristic acid. The probes bind to and, upon irradiation, covalently label the lipid-binding chaperone protein uncoordinated 119 (UNC119). UNC119 increases overall solubility and regulates specifically the transport of myristoylated proteins between intercellular membranes. The binding mode of the probes is similar to that of the myristate moiety, and the residues inside the hydrophobic pocket of UNC119 proteins that are critical for covalent binding have been identified. The interaction with UNC119 was also demonstrated in cell lysate by means of affinity enrichment. Moreover, it is shown that the myristate analogue can be incorporated into peptide substrates by N-myristoyl transferases of Leishmania and Trypanosoma protozoan parasites.
蛋白质豆蔻酰化在生物过程中起着关键作用，例如，在蛋白质的膜附着和激活以及介导蛋白质-蛋白质和蛋白质-脂质相互作用中。此外，豆蔻酰化蛋白与包括癌症和病毒感染在内的疾病有关。因此，研究蛋白质豆蔻酰化的新工具需求量很大。在此，我们报告了基于豆蔻酸的氮丙啶取代类似物开发的光活化探针。这些探针与结合，并在照射时，将脂质结合伴侣蛋白 UNC119 共价标记。UNC119 提高了整体溶解度，并特异性调节豆蔻酰化蛋白在细胞间膜之间的运输。探针的结合模式与豆蔻酸部分相似，并且鉴定出 UNC119 蛋白的疏水口袋内对于共价结合至关重要的残基。通过亲和富集在细胞裂解物中也证明了与 UNC119 的相互作用。此外，还表明可以通过利什曼原虫和原生动物寄生虫的 N-豆蔻酰转移酶将豆蔻酸类似物掺入肽底物中。
