Blue light-regulated molecular switch of Ser/Thr kinase in phototropin.
Phototropin is a blue light photoreceptor for tropic responses, relocation of chloroplasts, and stomata opening in plants. Phototropin has two chromophoric domains named light-oxygen-voltage-sensing (LOV) 1 and 2 in the N-terminal half, and a serine/threonine (Ser/Thr) protein kinase motif in the C-terminal half. Concerning the kinase activity of phototropin, only autophosphorylation has been detected so far. However, we found that phototropin can phosphorylate a protein other than phototropin itself. Bacterially expressed Arabidopsis phototropin 2 kinase domain (KD) with GST-tag showed a constitutive kinase activity on casein, a common in vitro substrate of Ser/Thr protein kinase. By using this in vitro assay system, the roles of each LOV domain were studied. Addition of LOV2 to KD (GST-L2-KD) inhibits the kinase activity that is canceled by light. This light activation of kinase disappeared on introduction of a mutation blocking photochemical reaction in the LOV2 domain. Accordingly, LOV2 domain acts as a major light-regulated molecular switch of casein phosphorylation. Interestingly, isolated LOV2 from the KD still binds to the KD in a light-dependent manner and functions in similar ways, indicating the role of LOV2 domain as an inhibitor of the kinase activity in the substrate phosphorylation. LOV1, in contrast, contributes little to the photoactivation in GST-L1-L2-KD; however, it acts as an attenuator of the light activation of the kinase by LOV2.
Matsuoka
Daisuke
D
Research Institute for Advanced Science and Technology, Osaka Prefecture University, 1-2 Gakuen-cho, Sakai, Osaka 599-8570, Japan.
Tokutomi
Satoru
S
D017360:Arabidopsis / Q000737:chemistry; D029681:Arabidopsis Proteins; D002364:Caseins / Q000378:metabolism; D056931:Cryptochromes; D005420:Flavoproteins / Q000737:chemistry / Q000378:metabolism*; D008027:Light; D010766:Phosphorylation; D010786:Photoreceptor Cells / Q000737:chemistry / Q000378:metabolism*; D010940:Plant Proteins; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D021122:Protein Subunits / Q000378:metabolism / Q000502:physiology
D029681:Arabidopsis Proteins; C538962:CRY1 protein, Arabidopsis; D002364:Caseins; D056931:Cryptochromes; D005420:Flavoproteins; D010940:Plant Proteins; D021122:Protein Subunits; D017346:Protein Serine-Threonine Kinases