Metal ion-modulated KRAS conformational dynamics unraveled by ultraviolet photodissociation.

Metal ions are indispensable cofactors governing protein conformations and biofunctions. However, given the highly dynamic and subtle nature of metal ion modulations, capturing the metal-induced conformational dynamics and resolving these structural alteration details remain challenging using traditional structural biology methods. Herein, we apply native mass spectrometry (nMS) and 193-nm ultraviolet photodissociation (UVPD) to characterize the metal ion-dependent conformations of Kirsten rat sarcoma (KRAS). Achieving high sequence coverage, we established a biophysical baseline for the native Mg2+ cofactor, demonstrating how its structural impact modulates functional switch loops to stabilize a compact and inactive state of KRAS. Furthermore, we find the competitive substitution of Mg2+ by Zn2+ preserves the global compact topology but alters local stability, imparting the active site and switch regions with improved flexibility. Our work highlights how metal ion modulates on KRAS conformational dynamics, providing a sensitive MS-based analytical strategy for exploring the allosteric modulations across broader metal-proteins.