title: A single circularly permuted GFP sensor for inositol-1,3,4,5-tetrakisphosphate based on a split PH domain.
abstract: A fluorescent sensor for the detection of inositol-1,3,4,5-tetrakisphosphate, Ins(1,3,4,5)P(4), was constructed from a split PH domain and a single circularly permuted GFP. A structure-based design was conducted to transduce a ligand-induced subtle structural perturbation of the split PH domain to an alteration in the population of the protonated and the deprotonated states of the GFP chromophore. Excitation of each distinct absorption band corresponding to the protonated or the deprotonated state of GFP resulted an increase and a decrease, respectively, in the intensity of emission spectra upon addition of Ins(1,3,4,5)P(4) to the split PH domain-based sensor. The Ins(1,3,4,5)P(4) sensor retained the ligand affinity and the selectivity of the parent PH domain, and realized the ratiometric fluorescence detection of Ins(1,3,4,5)P(4).
authors.affiliation: Institute of Advanced Energy, Kyoto University, Uji, Kyoto 611-0011, Japan.
authors.forename: Reiko
authors.initials: R
authors.lastname: Sakaguchi
authors.forename: Takashi
authors.initials: T
authors.lastname: Endoh
authors.forename: Seigo
authors.initials: S
authors.lastname: Yamamoto
authors.forename: Kazuki
authors.initials: K
authors.lastname: Tainaka
authors.forename: Kenji
authors.initials: K
authors.lastname: Sugimoto
authors.forename: Nobutaka
authors.initials: N
authors.lastname: Fujieda
authors.forename: Shigeki
authors.initials: S
authors.lastname: Kiyonaka
authors.forename: Yasuo
authors.initials: Y
authors.lastname: Mori
authors.forename: Takashi
authors.initials: T
authors.lastname: Morii
mesh_terms: D000595:Amino Acid Sequence; D001798:Blood Proteins / Q000737:chemistry; D031541:Fluorescence Resonance Energy Transfer; D049452:Green Fluorescent Proteins / Q000737:chemistry*; D007295:Inositol Phosphates; D008969:Molecular Sequence Data; D010750:Phosphoproteins / Q000737:chemistry; D011874:Radiometry
chemical_list: D001798:Blood Proteins; D007295:Inositol Phosphates; D010750:Phosphoproteins; C039077:platelet protein P47; C081655:inositol-1,3,4,5-tetrakisphosphate; D049452:Green Fluorescent Proteins