Overexpression in Yeast, Photocycle, and in Vitro Structural Change of an Avian Putative Magnetoreceptor Cryptochrome4

Hiromasa Mitsui, Toshinori Maeda, Chiaki Yamaguchi, Yusuke Tsuji, Ryuji Watari, Yoko Kubo, Keiko Okano, Toshiyuki Okano

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    Abstract

    Cryptochromes (CRYs) have been found in a wide variety of living organisms and can function as blue light photoreceptors, circadian clock molecules, or magnetoreceptors. Non-mammalian vertebrates have CRY4 in addition to the CRY1 and CRY2 circadian clock components. Though the function of CRY4 is not well understood, chicken CRY4 (cCRY4) may be a magnetoreceptor because of its high level of expression in the retina and light-dependent structural changes in retinal homogenates. To further characterize the photosensitive nature of cCRY4, we developed an expression system using budding yeast and purified cCRY4 at yields of submilligrams of protein per liter with binding of the flavin adenine dinucleotide (FAD) chromophore. Recombinant cCRY4 dissociated from anti-cCRY4 C1 mAb, which recognizes the C-terminal region of cCRY4, in a light-dependent manner and showed a light-dependent change in its trypsin digestion pattern, suggesting that cCRY4 changes its conformation with light irradiation in the absence of other retinal factors. Combinatorial analyses with UV - visible spectroscopy and immunoprecipitation revealed that there is chromophore reduction in the cCRY4 photocycle and formation of a flavosemiquinone radical intermediate that is likely accompanied by a conformational change in the carboxyl-terminal region. Thus, cCRY4 seems to be an intrinsically photosensitive and photoswitchable molecule and may exemplify a vertebrate model of cryptochrome with possible function as a photosensor and/or magnetoreceptor. (Figure Presented).

    Original languageEnglish
    Pages (from-to)1908-1917
    Number of pages10
    JournalBiochemistry
    Volume54
    Issue number10
    DOIs
    Publication statusPublished - 2015 Mar 17

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    ASJC Scopus subject areas

    • Biochemistry

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