Light-triggered conformational changes of an animal-like cryptochrome tracked by native time-resolved IMS-MS

Abstract Cryptochromes (CRYs) belong to the class of blue light photoreceptors and are responsible for various light-triggered functions in the circadian rhythm upon excitation of their inbuilt flavin cofactor. They are functionally distinct to the evolutionarily related photolyases, which mediate light-driven repair of UV induced DNA damages. Despite these functional differences they share a high degree of structural and sequence homology. A hallmark of cryptochromes is their flexible carboxyl-terminal extension (CTE), whose structure and function as well as the details of its interaction with the photolyase homology region (PHR) are not yet fully understood and differs among different cryptochromes types. In this study we investigate the animal-like cryptochrome from Chlamydomonas reinhardtii (Cr aCRY ). Here, we focus on the highly conserved C-terminal domain harboring the FAD chromophore, to study the effect of single mutations on the structural transition of the C-terminal helix α22 and the attached CTE upon lit-state formation. By coupling a high-power LED, mounted in the source region to an ion mobility mass spectrometer, we show that D321, the putative proton acceptor of the terminal proton-coupled electron transfer event from Y373, is essential for triggering the large-scale conformational changes of helix α22 and the CTE in the lit state.