| The phototropins are flavoprotein kinases that control phototropic bending, stomatal opening, and light-induced chloroplast movement in plants. Two flavin mononucleotide (FMN)-binding light, oxygen, or voltage (LOV) domains are the sites for initial photochemistry in these blue-light photoreceptors. The phototropin LOV domains form a subset of the large and diverse Per-ARNT-Sim (PAS) domain superfamily, which is more widely implicated in diverse signal transduction processes and in dimerization. This thesis describes the crystal structure determination of the dark and photoexcited forms of a FMN-bound LOV2 domain to 2.7 and 2.6 Å resolution, respectively. The LOV2 structure constitutes a novel FMN-binding fold that reveals how the flavin cofactor is embedded in the protein. In the dark structure, a single cysteine residue is located 4.2 Å from flavin atom C(4a), consistent with a model in which absorption of blue light induces formation of a covalent cysteinyl-C(4a) adduct. Indeed, the photoexcited structure reveals a unique photochemical switch in which absorption of light drives the formation of a reversible covalent bond between a highly conserved cysteine residue and C(4a) of the flavin cofactor. This provides the first molecular picture of a cysteinyl-flavin covalent adduct, the presumed signaling species that leads to phototropin kinase activation and subsequent signal transduction. Flavin-binding LOV domains, first identified in the higher plant phototropins, are shown to be present in other plant, fungal, and bacterial proteins. Notably, LOV domains are coupled to a wide array of other domains including kinases, phosphodiesterases, F-box domains, STAS domains, and zinc fingers, which suggests that the absorption of blue light by LOV regulates the activity of these structurally- and functionally-diverse domains. LOV domains contain a conserved molecular volume extending from the flavin cofactor, which is the locus for light-driven structural change, to the molecular surface. The role of this conserved volume of structure in LOV-regulated processes is discussed. |
