Discovering Targetable Conformation of RhoA Mutant by Integrating Native Mass Spectrometry, Ultraviolet Photodissociation, and X-ray Diffraction

Journal of the American Chemical Society 2026

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Authors: Haozhe Wu, Zheyi Liu, Haibo Jiang, Heng Zhao, Chen Dong, Yu Lu, Shijia Zu, Yongjie Guo, Can Lai, Pan Luo, Ke Xu, Yuxin Yang
+18 more Yafang Yang, Zhongya Sun, Qingcheng Huang, Huan Xiong, Lanxin Zhou, Yipan Luo, Yuhan Zeng, Daohai Du, Zhongjie Liang, Weilie Xiao, Shan Zhao, Weiqing Zhang, Yong Tang, Chunlei Xiao, Kaixian Chen, Xueming Yang, Fangjun Wang, Cheng Luo
DOI: 10.1021/jacs.5c20067
Cited by: 3
References: 40

Abstract

Pathogenic mutations in “undruggable” Ras superfamily proteins challenge drug development by inducing subtle, dynamic conformational changes. Here, we integrated X-ray crystallography with native mass spectrometry and ultraviolet photodissociation (nMS-UVPD) to reveal a cryptic conformation in the oncogenic Y42C mutant of RhoA. While crystallography alone resolved two ambiguous structures, nMS-UVPD determined the dominant conformation by directly mapping the mutant’s conformational dynamics, identifying an enhanced Mg 2+ -locked conformation. We explored the mechanism of mutation impairing GTP hydrolysis. This state unmasks a previously hidden, druggable pocket adjacent to Cys42, guiding our identification of a covalent inhibitor. Our integrated approach establishes a roadmap for targeting pathogenic protein mutants previously considered “undruggable” due to their highly dynamic nature.

Fields of Study & Subjects

Protein Kinase Regulation and GTPase Signaling Enzyme Structure and Function Advanced Proteomics Techniques and Applications

Publication details

Journal / source
Journal of the American Chemical Society
Publisher
American Chemical Society (ACS)
Volume
148
Issue
11
Pages
11709–11718
Publication date
16 March 2026
Language
EN