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Mechanistic Insights into Light-Gated Anion Channels from Guillardia Theta

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The most effective optogenetic tools available for neuronal silencing are the light-gated anion channels found in the cryptophyte alga Guillardia theta (GtACRs). GtACRs have the highest chloride-conductance among natural or synthetic anion-conducting channelrhodopsins. Molecular mechanisms of GtACRs are of great interest for understanding this exceptional conductance. The ion transport of GtACRs are intimately tied to their photochemical reaction mechanism in which the lifetimes of open channel photointermediates regulate the channel conductance. So far, no studies have been able to accurately pinpoint the photointermediate corresponding to the open channel state. In this thesis, the photoreactions of wild-type GtACR1 and variants are investigated through multichannel time-resolved absorption spectroscopy. The results presented here demonstrate the existence of isospectral photointermediates and identify a previously unknown photointermediate. Additional insights into the photomechanism are made and a new open channel photointermediate is proposed. This contribution to unraveling channelrhodopsin mechanisms will help elucidate the underpinnings of GtACRs unusually high conductance and facilitate improved rational design of molecular tools for optogenetic applications.

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This item is under embargo until October 24, 2024.