A bacteriophytochrome Pr/Pfr heterodimer studied through single-particle time-resolved cryo-electron microscopy

Phytochromes are dimeric photoreceptors found in bacteria, fungi, and plants that reversibly interconvert between a red-absorbing Pr state and a far-red-absorbing Pfr state. In bacteria, phytochromes (BphPs) regulate diverse responses through a two-component signaling pathway comprising a C-terminal histidine kinase (HK) and a response regulator. A previous cryo-EM study of the wild-type BphP from Stigmatella aurantiaca (SaBphP2) revealed a stable Pr/Pfr heterodimer in which the two protomers adopt distinct Pr and Pfr conformations. Here, using the Spotiton technique, we captured the same heterodimer by illuminating SaBphP2 particles on the cryo-EM grid and vitrifying them 10 ms later. Comparison with the Pr/Pr homodimer reveals a 180° rotation of the HK domain, driven by an unwinding of the coiled-coil helices that connect the photosensory core to the enzymatic domain. The large-scale reorientations provide mechanistic insight into light-triggered signal transduction mediated by bacterial phytochromes. The transition of bacterial phytochromes from the red-absorbing Pr state to the far-red-absorbing Pfr state may proceed through the formation of a Pr/Pfr heterodimer, as revealed by a recent single-particle cryo-EM study of a phytochrome from the myxobacterium Stigmatella aurantiaca. By illuminating this protein immediately before vitrification, the authors captured the Pr/Pfr heterodimer intermediate on a millisecond timescale and observed a large rotation of the C-terminal histidine kinase domain, a conformational change that is likely to regulate light-induced signal transduction.