title: Absorption and fluorescence spectroscopic characterization of cryptochrome 3 from Arabidopsis thaliana.
abstract: The blue light photoreceptor cryptochrome 3 (cry3) from Arabidopsis thaliana was characterized at room temperature in vitro in aqueous solution by optical absorption and emission spectroscopic studies. The protein non-covalently binds the chromophores flavin adenine dinucleotide (FAD) and N5,N10-methenyl-5,6,7,8-tetrahydrofolate (MTHF). In the dark-adapted state of cry3, the bound FAD is present in the oxidized form (FAD(ox), ca. 38.5%), in the semiquinone form (FADH., ca. 5%), and in the fully reduced neutral form (FAD(red)H2) or fully reduced anionic form (FAD(red)H-, ca. 55%). Some amount of FAD (ca. 1.5%) in the oxidized state remains unbound probably caused by chromophore release and/or denaturation. Förster-type energy transfer from MTHF to FAD(ox) is observed. Photo-excitation reversibly modifies the protein conformation causing a slight rise of the MTHF absorption strength and an increase of the MTHF fluorescence efficiency (efficient protein conformation photo-cycle). Additionally there occurs reversible reduction of bound FAD(ox) to FAD(red)H2 (or FAD(red)H-, FAD(ox) photo-cycle of moderate efficiency), reversible reduction of FADH. to FAD(red)H2 (or FAD(red)H-, FADH. photo-cycle of high efficiency), and modification of re-oxidable FAD(red)H2 (or FAD(red)H-) to permanent FAD(red)H2 (or FAD(red)H-) with low quantum efficiency. Photo-excitation of MTHF causes the reversible formation of a MTHF species (MTHF', MTHF photo-cycle, moderate quantum efficiency) with slow recovery to the initial dark state, and also the formation of an irreversible photoproduct (MTHF'').
authors.affiliation: Institut für Physikalische und Theoretische Chemie, Universität Regensburg, 93053 Regensburg, Germany.
authors.forename: S-H
authors.initials: SH
authors.lastname: Song
authors.forename: B
authors.initials: B
authors.lastname: Dick
authors.forename: A
authors.initials: A
authors.lastname: Penzkofer
authors.forename: R
authors.initials: R
authors.lastname: Pokorny
authors.forename: A
authors.initials: A
authors.lastname: Batschauer
authors.forename: L-O
authors.initials: LO
authors.lastname: Essen
mesh_terms: D000042:Absorption; D017360:Arabidopsis / Q000737:chemistry*; D029681:Arabidopsis Proteins / Q000032:analysis* / Q000737:chemistry / Q000378:metabolism; D056931:Cryptochromes; D004255:Deoxyribodipyrimidine Photo-Lyase / Q000032:analysis* / Q000737:chemistry / Q000378:metabolism; D004579:Electron Transport; D004735:Energy Transfer; D005182:Flavin-Adenine Dinucleotide / Q000737:chemistry / Q000378:metabolism*; D005420:Flavoproteins / Q000737:chemistry*; D005492:Folic Acid / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism; D011809:Quinones / Q000737:chemistry / Q000378:metabolism; D013050:Spectrometry, Fluorescence / Q000379:methods; D013696:Temperature; D013997:Time Factors
chemical_list: D029681:Arabidopsis Proteins; C538962:CRY1 protein, Arabidopsis; D056931:Cryptochromes; D005420:Flavoproteins; D011809:Quinones; D005182:Flavin-Adenine Dinucleotide; C032418:5,11-methenyltetrahydrohomofolate; D005492:Folic Acid; C515145:Cry3 protein, Arabidopsis; D004255:Deoxyribodipyrimidine Photo-Lyase