title: Essential role of the A'α/Aβ gap in the N-terminal upstream of LOV2 for the blue light signaling from LOV2 to kinase in Arabidopsis photototropin1, a plant blue light receptor.
abstract: Phototropin (phot) is a blue light (BL) receptor in plants and is involved in phototropism, chloroplast movement, stomata opening, etc. A phot molecule has two photo-receptive domains named LOV (Light-Oxygen-Voltage) 1 and 2 in its N-terminal region and a serine/threonine kinase (STK) in its C-terminal region. STK activity is regulated mainly by LOV2, which has a cyclic photoreaction, including the transient formation of a flavin mononucleotide (FMN)-cysteinyl adduct (S390). One of the key events for the propagation of the BL signal from LOV2 to STK is conformational changes in a Jα-helix residing downstream of the LOV2 C-terminus. In contrast, we focused on the role of the A'α-helix, which is located upstream of the LOV2 N-terminus and interacts with the Jα-helix. Using LOV2-STK polypeptides from Arabidopsis thaliana phot1, we found that truncation of the A'α-helix and amino acid substitutions at Glu474 and Lys475 in the gap between the A'α and the Aβ strand of LOV2 (A'α/Aβ gap) to Ala impaired the BL-induced activation of the STK, although they did not affect S390 formation. Trypsin digested the LOV2-STK at Lys603 and Lys475 in a light-dependent manner indicating BL-induced structural changes in both the Jα-helix and the gap. The digestion at Lys603 is faster than at Lys475. These BL-induced structural changes were observed with the Glu474Ala and the Lys475Ala substitutes, indicating that the BL signal reached the Jα-helix as well as the A'α/Aβ gap but could not activate STK. The amino acid residues, Glu474 and Lys475, in the gap are conserved among the phots of higher plants and may act as a joint to connect the structural changes in the Jα-helix with the activation of STK.
authors.affiliation: Department of Biological Science, Osaka Prefecture University, Sakai, Osaka, Japan.
authors.forename: Sachiko
authors.initials: S
authors.lastname: Kashojiya
authors.affiliation: Department of Biological Science, Osaka Prefecture University, Sakai, Osaka, Japan.
authors.forename: Koji
authors.initials: K
authors.lastname: Okajima
authors.affiliation: Life Science Research Center, SHIMADZU Corporation, Tokyo, Japan.
authors.forename: Takashi
authors.initials: T
authors.lastname: Shimada
authors.affiliation: Department of Biological Science, Osaka Prefecture University, Sakai, Osaka, Japan.
authors.forename: Satoru
authors.initials: S
authors.lastname: Tokutomi
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D017360:Arabidopsis / Q000235:genetics / Q000378:metabolism / Q000528:radiation effects*; D029681:Arabidopsis Proteins / Q000737:chemistry* / Q000235:genetics / Q000528:radiation effects; D003545:Cysteine / Q000737:chemistry; D004789:Enzyme Activation / Q000528:radiation effects; D005486:Flavin Mononucleotide / Q000378:metabolism; D006860:Hydrogen Bonding; D008027:Light; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010449:Peptide Mapping; D010750:Phosphoproteins / Q000737:chemistry* / Q000235:genetics / Q000528:radiation effects; D010766:Phosphorylation / Q000528:radiation effects; D010777:Photochemistry; D011487:Protein Conformation; D017346:Protein Serine-Threonine Kinases; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D016415:Sequence Alignment; D017384:Sequence Deletion; D017386:Sequence Homology, Amino Acid; D013329:Structure-Activity Relationship
chemical_list: D029681:Arabidopsis Proteins; D010750:Phosphoproteins; D005486:Flavin Mononucleotide; C103646:NPH1 protein, Arabidopsis; D017346:Protein Serine-Threonine Kinases; D003545:Cysteine