TY - JOUR
T1 - Implications for the Light-Driven Chloride Ion Transport Mechanism of Nonlabens marinus Rhodopsin 3 by Its Photochemical Characteristics
AU - Tsukamoto, Takashi
AU - Yoshizawa, Susumu
AU - Kikukawa, Takashi
AU - Demura, Makoto
AU - Sudo, Yuki
N1 - Funding Information:
This research was supported by grants from the Japanese Ministry of Education, Culture, Sports, Science, and Technology (MEXT) to T.T. (JP15K18519), to S.Y. (JP15H02800) and to Y.S. (JP15H04363). This work was also supported by a Grant-in-Aid for Scientific Research on Innovative Areas (25104005) and CREST, JST to Y.S. The authors thank DASS Manuscript for English language editing.
Publisher Copyright:
© 2017 American Chemical Society.
PY - 2017/3/9
Y1 - 2017/3/9
N2 - Several new retinal-based photoreceptor proteins that act as light-driven electrogenic halide ion pumps have recently been discovered. Some of them, called "NTQ" rhodopsins, contain a conserved Asn-Thr-Gln motif in the third or C-helix. In this study, we investigated the photochemical characteristics of an NTQ rhodopsin, Nonlabens marinus rhodopsin 3 (NM-R3), which was discovered in the N. marinus S1-08T strain, using static and time-resolved spectroscopic techniques. We demonstrate that NM-R3 binds a Cl- in the vicinity of the retinal chromophore accompanied by a spectral blueshift from 568 nm in the absence of Cl- to 534 nm in the presence of Cl-. From the Cl- concentration dependence, we estimated the affinity (dissociation constant, Kd) for Cl- in the original state as 24 mM, which is ca. 10 times weaker than that of archaeal halorhodopsins but ca. 3 times stronger than that of a marine bacterial Cl- pumping rhodopsin (C1R). NM-R3 showed no dark-light adaptation of the retinal chromophore and predominantly possessed an all-trans-retinal, which is responsible for the light-driven Cl- pump function. Flash-photolysis experiments suggest that NM-R3 passes through five or six photochemically distinct intermediates (K, L(N), O1, O2, and NM-R3′). From these results, we assume that the Cl- is released and taken up during the L(N)-O1 transition from a transiently formed cytoplasmic (CP) binding site and the O2-NM-R3′ or the NM-R3′-original NM-R3 transitions from the extracellular (EC) side, respectively. We propose a mechanism for the Cl- transport by NM-R3 based on our results and its recently reported crystal structure. (Chemical Equation Presented).
AB - Several new retinal-based photoreceptor proteins that act as light-driven electrogenic halide ion pumps have recently been discovered. Some of them, called "NTQ" rhodopsins, contain a conserved Asn-Thr-Gln motif in the third or C-helix. In this study, we investigated the photochemical characteristics of an NTQ rhodopsin, Nonlabens marinus rhodopsin 3 (NM-R3), which was discovered in the N. marinus S1-08T strain, using static and time-resolved spectroscopic techniques. We demonstrate that NM-R3 binds a Cl- in the vicinity of the retinal chromophore accompanied by a spectral blueshift from 568 nm in the absence of Cl- to 534 nm in the presence of Cl-. From the Cl- concentration dependence, we estimated the affinity (dissociation constant, Kd) for Cl- in the original state as 24 mM, which is ca. 10 times weaker than that of archaeal halorhodopsins but ca. 3 times stronger than that of a marine bacterial Cl- pumping rhodopsin (C1R). NM-R3 showed no dark-light adaptation of the retinal chromophore and predominantly possessed an all-trans-retinal, which is responsible for the light-driven Cl- pump function. Flash-photolysis experiments suggest that NM-R3 passes through five or six photochemically distinct intermediates (K, L(N), O1, O2, and NM-R3′). From these results, we assume that the Cl- is released and taken up during the L(N)-O1 transition from a transiently formed cytoplasmic (CP) binding site and the O2-NM-R3′ or the NM-R3′-original NM-R3 transitions from the extracellular (EC) side, respectively. We propose a mechanism for the Cl- transport by NM-R3 based on our results and its recently reported crystal structure. (Chemical Equation Presented).
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U2 - 10.1021/acs.jpcb.6b11101
DO - 10.1021/acs.jpcb.6b11101
M3 - Article
C2 - 28194973
AN - SCOPUS:85015913111
VL - 121
SP - 2027
EP - 2038
JO - Journal of Physical Chemistry B
JF - Journal of Physical Chemistry B
SN - 1520-6106
IS - 9
ER -