Author:

Keywords:

photoactivation-localization microscopy, crystal-structure, peptide cleavage, structural basis, resolution, coral, eosfp, gfp, Science & Technology, Life Sciences & Biomedicine, Biochemistry & Molecular Biology, CRYSTAL-STRUCTURE, PEPTIDE CLEAVAGE, STRUCTURAL BASIS, MICROSCOPY, CORAL, RESOLUTION, GFP, Amino Acid Sequence, Crystallography, X-Ray, Luminescent Proteins, Molecular Sequence Data, Photochemical Processes, Protein Conformation, Recombinant Proteins, Sequence Alignment, Ultraviolet Rays, 0304 Medicinal and Biomolecular Chemistry, 0305 Organic Chemistry, 0601 Biochemistry and Cell Biology, Organic Chemistry, 3101 Biochemistry and cell biology, 3404 Medicinal and biomolecular chemistry

Abstract:

KikGR is a fluorescent protein engineered to display green-to-red photoconvertibility that is induced by irradiation with ultraviolet or violet light. Similar to Kaede and EosFP, two naturally occurring photoconvertible proteins, KikGR contains a HiS(62)-Tyr(63)-Gly(64) tripeptide sequence, which forms a green chromophore that can be photoconverted to a red one via formal beta-elimination and subsequent extension of a pi-conjugated system. Using a crystallizable variant of KikGR, we determined the structures of both the green and red state at 1.55 angstrom. resolution. The double bond between HiS(62)-C-alpha and HiS(62)-C-beta in the red chromophore is in a cis configuration, indicating that rotation along the HiS(62) C-alpha-C-beta bond occurs following cleavage of the HiS(62) N-alpha-C-alpha bond. This structural rearrangement provides evidence that the beta-elimination reaction governing the green-to-red photoconversion of KikGR follows an E1 (elimination, unimolecular) mechanism.