Combined Mutational and Spectroscopic Study on the Calcium-Related Kinetic Effects on the VirChR1 Photocycle

Gerrit H. U. Lamm; Dmitrii Zabelskii; Taras Balandin; Valentin Gordeliy; Josef Wachtveitl

doi:10.1021/acs.jpcb.4c08416

Combined Mutational and Spectroscopic Study on the Calcium-Related Kinetic Effects on the VirChR1 Photocycle

Gerrit H. U. Lamm, Dmitrii Zabelskii, Taras Balandin, Valentin Gordeliy, Josef Wachtveitl

Research output: Contribution to Journal › Article › Academic › peer-review

Abstract

The viral rhodopsin 1 subfamily consists of microbial rhodopsins, such as VirChR1, with a light-gated cation channeling functionality, which is inhibited by calcium. For VirChR1, S14, E54, and N225 have been proposed as key residues for calcium binding. They form a highly conserved SEN-triad in channelrhodopsins near the functionally important central gate. Here, we present a time-resolved UV/vis spectroscopic study on the VirChR1 variants S14A, E54A, and N225A in a calcium-dependent manner. Comparison with the calcium-associated effects observed for the wild type shed light on the role of the respective residues for the calcium interaction. While S14A shows less pronounced, yet similar, signals, indicative of a reduced calcium affinity, E54A exhibits nearly calcium-independent photocycle kinetics, highlighting its crucial role for calcium binding. The N225A variant shows altered photocycle kinetics, in both the absence and presence of calcium, demonstrating its critical role in the formation of the functionally important central gate in VirChR1.

Original language	English
Pages (from-to)	2946-2957
Journal	Journal of Physical Chemistry B
Volume	129
Issue number	11
DOIs	https://doi.org/10.1021/acs.jpcb.4c08416
Publication status	Published - 20 Mar 2025
Externally published	Yes

Funding

This research was supported by the German Research Foundation (CRC 1507\u2500Membrane-associated Protein Assemblies, Machineries, and Supercomplexes; Project P05 to J.W.). Additionally, G.H.U.L. and J.W. thank Dr. Marvin Asido, Dr. Christian Bamann, and Dr. Kirill Kovalev for helpful discussions. Additionally, Dr. Kirill Kovalev is further acknowledged for helping with the prediction of the VirChR1 structure.

Funders	Funder number
Deutsche Forschungsgemeinschaft	CRC 1507

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title = "Combined Mutational and Spectroscopic Study on the Calcium-Related Kinetic Effects on the VirChR1 Photocycle",

abstract = "The viral rhodopsin 1 subfamily consists of microbial rhodopsins, such as VirChR1, with a light-gated cation channeling functionality, which is inhibited by calcium. For VirChR1, S14, E54, and N225 have been proposed as key residues for calcium binding. They form a highly conserved SEN-triad in channelrhodopsins near the functionally important central gate. Here, we present a time-resolved UV/vis spectroscopic study on the VirChR1 variants S14A, E54A, and N225A in a calcium-dependent manner. Comparison with the calcium-associated effects observed for the wild type shed light on the role of the respective residues for the calcium interaction. While S14A shows less pronounced, yet similar, signals, indicative of a reduced calcium affinity, E54A exhibits nearly calcium-independent photocycle kinetics, highlighting its crucial role for calcium binding. The N225A variant shows altered photocycle kinetics, in both the absence and presence of calcium, demonstrating its critical role in the formation of the functionally important central gate in VirChR1.",

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T1 - Combined Mutational and Spectroscopic Study on the Calcium-Related Kinetic Effects on the VirChR1 Photocycle

AU - Lamm, Gerrit H. U.

AU - Zabelskii, Dmitrii

AU - Balandin, Taras

AU - Gordeliy, Valentin

AU - Wachtveitl, Josef

PY - 2025/3/20

Y1 - 2025/3/20

N2 - The viral rhodopsin 1 subfamily consists of microbial rhodopsins, such as VirChR1, with a light-gated cation channeling functionality, which is inhibited by calcium. For VirChR1, S14, E54, and N225 have been proposed as key residues for calcium binding. They form a highly conserved SEN-triad in channelrhodopsins near the functionally important central gate. Here, we present a time-resolved UV/vis spectroscopic study on the VirChR1 variants S14A, E54A, and N225A in a calcium-dependent manner. Comparison with the calcium-associated effects observed for the wild type shed light on the role of the respective residues for the calcium interaction. While S14A shows less pronounced, yet similar, signals, indicative of a reduced calcium affinity, E54A exhibits nearly calcium-independent photocycle kinetics, highlighting its crucial role for calcium binding. The N225A variant shows altered photocycle kinetics, in both the absence and presence of calcium, demonstrating its critical role in the formation of the functionally important central gate in VirChR1.

AB - The viral rhodopsin 1 subfamily consists of microbial rhodopsins, such as VirChR1, with a light-gated cation channeling functionality, which is inhibited by calcium. For VirChR1, S14, E54, and N225 have been proposed as key residues for calcium binding. They form a highly conserved SEN-triad in channelrhodopsins near the functionally important central gate. Here, we present a time-resolved UV/vis spectroscopic study on the VirChR1 variants S14A, E54A, and N225A in a calcium-dependent manner. Comparison with the calcium-associated effects observed for the wild type shed light on the role of the respective residues for the calcium interaction. While S14A shows less pronounced, yet similar, signals, indicative of a reduced calcium affinity, E54A exhibits nearly calcium-independent photocycle kinetics, highlighting its crucial role for calcium binding. The N225A variant shows altered photocycle kinetics, in both the absence and presence of calcium, demonstrating its critical role in the formation of the functionally important central gate in VirChR1.

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U2 - 10.1021/acs.jpcb.4c08416

DO - 10.1021/acs.jpcb.4c08416

M3 - Article

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SP - 2946

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JO - Journal of Physical Chemistry B

JF - Journal of Physical Chemistry B

IS - 11

ER -

Combined Mutational and Spectroscopic Study on the Calcium-Related Kinetic Effects on the VirChR1 Photocycle

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