Towards visualizing the B12-dependent photoreceptor CarH in action with time-resolved cryo-electron microscopy

CarH is a tetrameric transcription factor that controls carotenoid production in bacteria in response to light by using vitamin B12 as a chromophore. Illumination triggers a series of chemical reactions in the protein that end in dissociation of the tetramer. CarH has potential applications in optogenetics, but a detailed understanding of its mechanism is still lacking. We will use time-resolved cryo-electron microscopy, which will allow us to obtain high-resolution structural information about the later steps of the photoreaction. The protein sample will be illuminated directly on a cryo-EM grid before being vitrified after a controlled time delay, in order to capture reaction intermediates. We have expressed and purified CarH, in full-length in complex with DNA fragments and prepared cryo-EM grids.