Abstract
Phototropin (phot), a blue light receptor for phototropism, chloroplast movement, stomatal opening, etc. has two LOV domains and Ser/Thr kinase in its N- and C-terminal regions, respectively. In the dark, LOV2 acts as an inhibitor of the kinase. Blue light initiates photoreaction including a transient covalent bond formation between a Cys and a bound FMN in the LOV, which is supposed to cancel the inhibition via structural changes in the Jαhelix in the linker region with the kinase. Based on the predicted 3D model of Arabidopsis phot1 LOV2, we presumed two intradomain signaling pathways from FMN to Jαhelix (1)FMN (isoalloxazine)-N476-K475 and (2)FMN (phosphate)-R529-E537). In order to examine these two pathways, these amino acids were mutated to Ala and spectroscopic measurement, peptide mapping and kinase assay were performed. Amino acid mutations in the (1) little affected on the spectroscopic properties and peptide mapping patterns, but abolished the light-induced kinase activation. In contrast, those in (2) showed the light activation proposing the involvement of path (1) in the intradomain signaling responsible for the light-induced activation of the phot1 kinase.
