Institute of Molecular Biology and Biophysics, ETH Zürich, Otto-Stern-Weg 5, 8093 Zürich (Switzerland).
Angew Chem Int Ed Engl. 2014 Jan 27;53(5):1320-3. doi: 10.1002/anie.201306702. Epub 2013 Dec 16.
Ste5 is a scaffold protein that controls the pheromone response of the MAP-kinase cascade in yeast cells. Upon pheromone stimulation, Ste5 (through its RING-H2 domain) interacts with the β and γ subunits of an activated heterodimeric G protein and promotes activation of the MAP-kinase cascade. With structural and biophysical studies, we show that the Ste5 RING-H2 domain exists as a molten globule under native buffer conditions, in yeast extracts, and even in denaturing conditions containing urea (7 M). Furthermore, it exhibits high thermal stability in native conditions. Binding of the Ste5 RING-H2 domain to the physiological Gβ/γ (Ste4/Ste18) ligand is accompanied by a conformational transition into a better folded, more globular structure. This study reveals novel insights into the folding mechanism and recruitment of binding partners by the Ste5 RING-H2 domain. We speculate that many RING domains may share a similar mechanism of substrate recognition and molten-globule-like character.
Ste5 是一种支架蛋白，它控制着酵母细胞中 MAP 激酶级联的信息素反应。在信息素刺激下，Ste5（通过其 RING-H2 结构域）与激活的异源二聚体 G 蛋白的β和γ亚基相互作用，并促进 MAP 激酶级联的激活。通过结构和生物物理研究，我们表明 Ste5 RING-H2 结构域在天然缓冲条件下、酵母提取物中甚至在含有尿素（7M）的变性条件下都以无定形球蛋白的形式存在。此外，它在天然条件下表现出很高的热稳定性。Ste5 RING-H2 结构域与生理 Gβ/γ（Ste4/Ste18）配体的结合伴随着构象转变为更好折叠、更球形的结构。这项研究揭示了 Ste5 RING-H2 结构域折叠机制和结合伙伴招募的新见解。我们推测，许多 RING 结构域可能具有类似的底物识别机制和无定形球蛋白样特征。
