期刊
BIOCHIMICA ET BIOPHYSICA ACTA-GENERAL SUBJECTS
卷 1810, 期 7, 页码 675-682出版社
ELSEVIER SCIENCE BV
DOI: 10.1016/j.bbagen.2011.04.008
关键词
Ostreococcus tauri; Marine algae; Phototropin; LOV domain; Dimerization; Dark recovery kinetics
资金
- DST
Background: Phototropins are UV-A/blue light receptor proteins with two LOV (Light-Oxygen-Voltage) sensor domains at their N terminus and a kinase domain at the C-terminus in photoautotrophic organisms. This is the first research report of a canonical phototropin from marine algae Ostreococcus tauri. Methods: We synthesized core LOV1 (OtLOV1) domain-encoding portion of the phototropin gene of O. tauri, the domain was heterologously expressed, purified and assessed for its spectral properties and dark recovery kinetics by UV-Visible, fluorescence spectroscopy and mutational studies. Quaternary structure characteristics were studied by SEC and glutaraldehyde crosslinking. Results: The absorption spectrum of OtLOV1 lacks the characteristic 361 nm peak shown by other LOV1 domains. It undergoes a photocycle with a dark state recovery time of approximately 30 min (tau=300.35 s). Native OtLOV1 stayed as dimer in aqueous solution and the dimer formation was light and concentration independent. Mutating isoleucine at 43rd position to valine accelerated the dark recovery time by more than 10-fold. Mutating it to serine reduced sensitivity to blue light, but the dark recovery time remained unaltered. I43S mutation also destabilized the FMN binding to a great extent. Conclusion: The OtLOV1 domain of the newly identified OtPhot is functional and the isoleucine at position 43 of OtLOV1 is the key residue responsible for fine-tuning the domain properties. General significance: This is the first characterized LOV1 domain of a canonical phototropin from a marine alga and spectral properties of the domain are similar to that of the LOV1 domain of higher plants. (C) 2011 Elsevier B.V. All rights reserved.
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