Light modulation of histidine-kinase activity in bacterial phytochromes monitored by size exclusion chromatography, crosslinking, and limited proteolysis.

Phytochromes are photoreceptors that have been found in plants, bacteria, and fungi. Most bacterial and fungal phytochromes are histidine kinases and, for several bacterial phytochromes, light regulation of kinase activity has been demonstrated. Typical histidine kinases are homodimeric proteins in which one subunit phosphorylates the substrate histidine residue of the other subunit; dimerization is an intrinsic property of the histidine kinase itself.

Protein conformational changes of Agrobacterium phytochrome Agp1 during chromophore assembly and photoconversion.

Phytochromes are widely distributed photochromic biliprotein photoreceptors. Typical bacterial phytochromes such as Agrobacterium Agp1 have a C-terminal histidine kinase module; the N-terminal chromophore module induces conformational changes in the protein that lead to modulation of kinase activity. We show by protein cross-linking that the C-terminal histidine kinase module of Agp1 mediates stable dimerization.

Assembly of synthetic locked chromophores with agrobacterium phytochromes Agp1 and Agp2.

Phytochromes are photoreceptors with a bilin chromophore in which light triggers the conversion between the red-absorbing form Pr and the far-red-absorbing form Pfr. Agrobacterium tumefaciens has two phytochromes, Agp1 and Agp2, with antagonistic properties: in darkness, Agp1 converts slowly from Pfr to Pr, whereas Agp2 converts slowly from Pr to Pfr. In a previous study, we have assembled Agp1 with synthetic locked chromophores 15Za, 15Zs, 15Ea, and 15Es in which the C15=C16 double bond is fixed in either the E or Z configuration and the C14-C15 single bond is fixed in either the syn (s) or anti (a) conformation.

Crystallization and preliminary X-ray crystallographic analysis of the N-terminal photosensory module of phytochrome Agp1, a biliverdin-binding photoreceptor from Agrobacterium tumefaciens.

Phytochromes are photochromic photoreceptors with a bilin chromophore that have been found in plants and bacteria. Typical bacterial phytochromes are composed of an N-terminal photosensory chromophore module and a C-terminal protein kinase. The former contains the chromophore, which allows phytochromes to adopt the two interconvertible spectral forms, Pr and Pfr.

Sterically locked synthetic bilin derivatives and phytochrome Agp1 from Agrobacterium tumefaciens form photoinsensitive Pr- and Pfr-like adducts.

Phytochrome photoreceptors undergo reversible photoconversion between the red-absorbing form, Pr, and the far-red-absorbing form, Pfr. The first step in the conversion from Pr to Pfr is a Z to E isomerization around the C15=C16 double bond of the bilin chromophore. We prepared four synthetic biliverdin (BV) derivatives in which rings C and D are sterically locked by cyclizing with an additional carbon chain.