Characterization of photo-intermediates in the photo-reaction pathways of a bacteriorhodopsin Y185F mutant using in situ photo-irradiation solid-state NMR spectroscopy

Kyosuke Oshima, Arisu Shigeta, Yoshiteru Makino, Izuru Kawamura, Takashi Okitsu, Akimori Wada, Satoru Tuzi, Tatsuo Iwasa, Akira Naito

    Abstract

    13C CP MAS NMR spectra were recorded at -40 °C in the dark (D1), under irradiation with 520 nm light (L1), subsequently in the dark (D2), and again under irradiation with 520 nm light (L2). In the process from D1 to L1, the 13-cis, 15-syn (CS; bR548) state changed to a CS∗- (13-cis, 15-syn) intermediate, which was highly stable at -40 °C, and the all-trans (AT; bR568) state transformed to an N-intermediate. Under the D2 conditions, the N-intermediate transformed to an O-intermediate, which was highly stable at -40 °C in the dark. During subsequent irradiation with 520 nm light (L2), the O-intermediate transformed to the N-intermediate through the AT state, whereas the CS∗-intermediate did not change. The CS∗-intermediate was converted to the AT state (or O-intermediate) after the temperature was increased to -20 °C. Upon subsequent increase of the temperature to 20 °C, the AT state (or O-intermediate) was converted to the CS state until reaching equilibrium. In this experiment, the chemical shift values of [20-13C, 14-13C]retinal provided the 13CC and 15CN configurations, respectively. From these data, the configurations of the AT and CS states and the CS∗-, N-, and O-intermediates were determined to be (13-trans, 15-anti), (13-cis, 15-syn), (13-cis, 15-syn), (13-cis, 15-anti), and (13-trans, 15-anti), respectively. 13C NMR signals of the CS∗- and O-intermediates were observed for the first time for the Y185F bR mutant by in situ photo-irradiation solid-state NMR spectroscopy and the configuration of the CS∗-intermediate was revealed to be significantly twisted from that of the CS state although both were assigned as (13-cis, 15-syn) configurations.]]>

    Original languageEnglish
    Pages (from-to)1694-1702
    Number of pages9
    JournalPhotochemical and Photobiological Sciences
    Volume14
    Issue number9
    DOIs
    StatePublished - 2015 Jun 26

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    Bacteriorhodopsins
    Magnetic Resonance Spectroscopy
    Temperature

    ASJC Scopus subject areas

    • Physical and Theoretical Chemistry

    Cite this

    Characterization of photo-intermediates in the photo-reaction pathways of a bacteriorhodopsin Y185F mutant using in situ photo-irradiation solid-state NMR spectroscopy. / Oshima, Kyosuke; Shigeta, Arisu; Makino, Yoshiteru; Kawamura, Izuru; Okitsu, Takashi; Wada, Akimori; Tuzi, Satoru; Iwasa, Tatsuo; Naito, Akira.

    In: Photochemical and Photobiological Sciences, Vol. 14, No. 9, 26.06.2015, p. 1694-1702.

    Research output: Contribution to journalArticle

    Oshima, Kyosuke; Shigeta, Arisu; Makino, Yoshiteru; Kawamura, Izuru; Okitsu, Takashi; Wada, Akimori; Tuzi, Satoru; Iwasa, Tatsuo; Naito, Akira / Characterization of photo-intermediates in the photo-reaction pathways of a bacteriorhodopsin Y185F mutant using in situ photo-irradiation solid-state NMR spectroscopy.

    In: Photochemical and Photobiological Sciences, Vol. 14, No. 9, 26.06.2015, p. 1694-1702.

    Research output: Contribution to journalArticle

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    author = "Kyosuke Oshima and Arisu Shigeta and Yoshiteru Makino and Izuru Kawamura and Takashi Okitsu and Akimori Wada and Satoru Tuzi and Tatsuo Iwasa and Akira Naito",
    year = "2015",
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    T1 - Characterization of photo-intermediates in the photo-reaction pathways of a bacteriorhodopsin Y185F mutant using in situ photo-irradiation solid-state NMR spectroscopy

    AU - Oshima,Kyosuke

    AU - Shigeta,Arisu

    AU - Makino,Yoshiteru

    AU - Kawamura,Izuru

    AU - Okitsu,Takashi

    AU - Wada,Akimori

    AU - Tuzi,Satoru

    AU - Iwasa,Tatsuo

    AU - Naito,Akira

    PY - 2015/6/26

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    N2 - Photo-reaction pathways of a bacteriorhodopsin Y185F mutant were examined using in situ photo-irradiation solid-state NMR spectroscopy. 13C CP MAS NMR spectra were recorded at -40 °C in the dark (D1), under irradiation with 520 nm light (L1), subsequently in the dark (D2), and again under irradiation with 520 nm light (L2). In the process from D1 to L1, the 13-cis, 15-syn (CS; bR548) state changed to a CS∗- (13-cis, 15-syn) intermediate, which was highly stable at -40 °C, and the all-trans (AT; bR568) state transformed to an N-intermediate. Under the D2 conditions, the N-intermediate transformed to an O-intermediate, which was highly stable at -40 °C in the dark. During subsequent irradiation with 520 nm light (L2), the O-intermediate transformed to the N-intermediate through the AT state, whereas the CS∗-intermediate did not change. The CS∗-intermediate was converted to the AT state (or O-intermediate) after the temperature was increased to -20 °C. Upon subsequent increase of the temperature to 20 °C, the AT state (or O-intermediate) was converted to the CS state until reaching equilibrium. In this experiment, the chemical shift values of [20-13C, 14-13C]retinal provided the 13CC and 15CN configurations, respectively. From these data, the configurations of the AT and CS states and the CS∗-, N-, and O-intermediates were determined to be (13-trans, 15-anti), (13-cis, 15-syn), (13-cis, 15-syn), (13-cis, 15-anti), and (13-trans, 15-anti), respectively. 13C NMR signals of the CS∗- and O-intermediates were observed for the first time for the Y185F bR mutant by in situ photo-irradiation solid-state NMR spectroscopy and the configuration of the CS∗-intermediate was revealed to be significantly twisted from that of the CS state although both were assigned as (13-cis, 15-syn) configurations.

    AB - Photo-reaction pathways of a bacteriorhodopsin Y185F mutant were examined using in situ photo-irradiation solid-state NMR spectroscopy. 13C CP MAS NMR spectra were recorded at -40 °C in the dark (D1), under irradiation with 520 nm light (L1), subsequently in the dark (D2), and again under irradiation with 520 nm light (L2). In the process from D1 to L1, the 13-cis, 15-syn (CS; bR548) state changed to a CS∗- (13-cis, 15-syn) intermediate, which was highly stable at -40 °C, and the all-trans (AT; bR568) state transformed to an N-intermediate. Under the D2 conditions, the N-intermediate transformed to an O-intermediate, which was highly stable at -40 °C in the dark. During subsequent irradiation with 520 nm light (L2), the O-intermediate transformed to the N-intermediate through the AT state, whereas the CS∗-intermediate did not change. The CS∗-intermediate was converted to the AT state (or O-intermediate) after the temperature was increased to -20 °C. Upon subsequent increase of the temperature to 20 °C, the AT state (or O-intermediate) was converted to the CS state until reaching equilibrium. In this experiment, the chemical shift values of [20-13C, 14-13C]retinal provided the 13CC and 15CN configurations, respectively. From these data, the configurations of the AT and CS states and the CS∗-, N-, and O-intermediates were determined to be (13-trans, 15-anti), (13-cis, 15-syn), (13-cis, 15-syn), (13-cis, 15-anti), and (13-trans, 15-anti), respectively. 13C NMR signals of the CS∗- and O-intermediates were observed for the first time for the Y185F bR mutant by in situ photo-irradiation solid-state NMR spectroscopy and the configuration of the CS∗-intermediate was revealed to be significantly twisted from that of the CS state although both were assigned as (13-cis, 15-syn) configurations.

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