Department of Chemistry, Middle Tennessee State University, 1301 East Main Street, Murfreesboro, TN 37132, USA.
Department of Chemistry, Tennessee Tech University, 1 William L Jones Drive, Cookeville, TN 38505, USA.
Int J Mol Sci. 2018 Nov 19;19(11):3651. doi: 10.3390/ijms19113651.
ClpC1 hexamers couple the energy of ATP hydrolysis to unfold and, subsequently, translocate specific protein substrates into the associated ClpP protease. Substrate recognition by ATPases associated with various cellular activities (AAA+) proteases is driven by the ATPase component, which selectively determines protein substrates to be degraded. The specificity of these unfoldases for protein substrates is often controlled by an adaptor protein with examples that include MecA regulation of ClpC or ClpS-mediated control of ClpA. No adaptor protein-mediated control has been reported for mycobacterial ClpC1. Using pulldown and stopped-flow fluorescence methods, we report data demonstrating that ClpC1 catalyzed unfolding of an SsrA-tagged protein is negatively impacted by association with the ClpS adaptor protein. Our data indicate that ClpS-dependent inhibition of ClpC1 catalyzed SsrA-dependent protein unfolding does not require the ClpC1 N-terminal domain but instead requires the presence of an interaction surface located in the ClpC1 Middle Domain. Taken together, our results demonstrate for the first time that mycobacterial ClpC1 is subject to adaptor protein-mediated regulation in vitro.
ClpC1 六聚体将 ATP 水解的能量耦合起来，展开并随后将特定的蛋白质底物转运到相关的 ClpP 蛋白酶中。与各种细胞活动相关的 ATP 酶（AAA+）蛋白酶对底物的识别是由 ATP 酶组件驱动的，该组件选择性地确定要降解的蛋白质底物。这些解折叠酶对蛋白质底物的特异性通常受衔接蛋白的控制，例如 MecA 对 ClpC 的调节或 ClpS 对 ClpA 的调节。尚未报道分枝杆菌 ClpC1 的衔接蛋白介导的控制。使用下拉和停流荧光方法，我们报告的数据表明，ClpS 衔接蛋白的结合会负调控 ClpC1 催化的 SsrA 标记蛋白的展开。我们的数据表明，ClpS 依赖性抑制 ClpC1 催化的 SsrA 依赖性蛋白展开不需要 ClpC1 N 端结构域，而是需要位于 ClpC1 中域的相互作用表面的存在。总之，我们的结果首次表明，分枝杆菌 ClpC1 在体外受到衔接蛋白介导的调控。
