ETH Zürich, Department of Biosystems Science and Engineering, Basel, Switzerland.
Proteomics. 2010 Dec;10(23):4151-62. doi: 10.1002/pmic.201000241.
We applied dynamic single-molecule force spectroscopy to quantify the parameters (free energy of activation and distance of the transition state from the folded state) characterizing the energy barriers in the unfolding energy landscape of the outer membrane protein G (OmpG) from Escherichia coli. The pH-dependent functional switching of OmpG directs the protein along different regions on the unfolding energy landscape. The two functional states of OmpG take the same unfolding pathway during the sequential unfolding of β-hairpins I-IV. After the initial unfolding events, the unfolding pathways diverge. In the open state, the unfolding of β-hairpin V in one step precedes the unfolding of β-hairpin VI. In the closed state, β-hairpin V and β-strand S11 with a part of extracellular loop L6 unfold cooperatively, and subsequently β-strand S12 unfolds with the remaining loop L6. These two unfolding pathways in the open and closed states join again in the last unfolding step of β-hairpin VII. Also, the conformational change from the open to the closed state witnesses a rigidified extracellular gating loop L6. Thus, a change in the conformational state of OmpG not only bifurcates its unfolding pathways but also tunes its mechanical properties for optimum function.
我们应用动态单分子力谱技术来量化描述大肠杆菌外膜蛋白 G（OmpG）解折叠能谱中能量势垒的参数（活化自由能和过渡态与折叠态的距离）。OmpG 的 pH 依赖性功能切换指导蛋白质沿着解折叠能谱上的不同区域运动。OmpG 的两种功能状态在β发夹 I-IV 的顺序解折叠过程中采用相同的解折叠途径。初始解折叠事件之后，解折叠途径开始分叉。在开放状态下，β发夹 V 一步解折叠先于β发夹 VI 解折叠。在闭合状态下，β发夹 V 和带有部分细胞外环 L6 的β-链 S11 协同解折叠，随后剩余的环 L6 与β-链 S12 一起解折叠。这两种在开放和闭合状态下的解折叠途径在β发夹 VII 的最后一步再次结合。此外，从开放状态到闭合状态的构象变化见证了刚性化的细胞外环 L6。因此，OmpG 构象状态的变化不仅使其解折叠途径分叉，而且还调节其机械性能以实现最佳功能。
