Prakash Ajit, Li Zijian, Chirasani Venkat R, Rasquinha Juhi A, Hewitt Natalie, Hubbard Garrett B, Yin Guowei, Hawkins Aspen T, Montore Luca J, Dohlman Henrik G, Campbell Sharon L
Department of Biochemistry & Biophysics, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA.
R. L. Juliano Structural Bioinformatics Core, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA.
Nat Commun. 2025 Apr 11;16(1):3468. doi: 10.1038/s41467-025-58323-2.
Heterotrimeric G proteins (Gα, Gβ and Gγ) act downstream of G-protein-coupled receptors (GPCRs) to mediate signaling pathways that regulate various physiological processes and human disease conditions. While human Gαi and its yeast homolog Gpa1 were previously postulated to function as intracellular pH sensors, the pH-sensing capabilities of Gαi and the underlying mechanism remain to be established. Our research shows that variations in pH significantly affect the structure and stability of Gαi-GDP. Specifically, at the lower end of the physiological pH range, the protein undergoes an order-to-disorder transition due to the loss of electrostatic interactions within the Gαi Switch regions, resulting in a reduction in agonist-mediated Gαi-Gβγ release. Further, we identified key residues within the Gαi Switch regions that form the pH-sensing network. Mutation of these residues in Gαi gives rise to 'low pH mimetics' that abolish pH-dependent thermostability changes and reduce Gαi-Gβγ release. Overall, our findings suggest that pH-sensitive structural changes in Gαi impact the agonist-mediated dissociation of Gβγ, which is essential for proper signaling.
异源三聚体G蛋白（Gα、Gβ和Gγ）在G蛋白偶联受体（GPCR）下游发挥作用，介导调节各种生理过程和人类疾病状态的信号通路。虽然人类Gαi及其酵母同源物Gpa1此前被假定为细胞内pH传感器，但Gαi的pH传感能力及其潜在机制仍有待确定。我们的研究表明，pH值的变化会显著影响Gαi-GDP的结构和稳定性。具体而言，在生理pH范围的下限，由于Gαi开关区域内静电相互作用的丧失，该蛋白会发生从有序到无序的转变，导致激动剂介导的Gαi-Gβγ释放减少。此外，我们确定了Gαi开关区域内形成pH传感网络的关键残基。Gαi中这些残基的突变产生了“低pH模拟物”，消除了pH依赖性热稳定性变化并减少了Gαi-Gβγ释放。总体而言，我们的研究结果表明，Gαi中对pH敏感的结构变化会影响激动剂介导的Gβγ解离，而这对于正确的信号传导至关重要。
