An Optogenetic Tool for Induced Protein Stabilization Based on the Phaeodactylum tricornutum Aureochrome 1a Light-Oxygen-Voltage Domain.,Journal of Molecular Biology

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An Optogenetic Tool for Induced Protein Stabilization Based on the Phaeodactylum tricornutum Aureochrome 1a Light-Oxygen-Voltage Domain.
Journal of Molecular Biology ( IF 4.7 ) Pub Date : 2020-02-24 , DOI: 10.1016/j.jmb.2020.02.019
Sebastian Hepp ₁ , Jonathan Trauth ₂ , Sophia Hasenjäger ₃ , Filipp Bezold ₁ , Lars-Oliver Essen ₁ , Christof Taxis ₃

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Control of cellular events by optogenetic tools is a powerful approach to manipulate cellular functions in a minimally invasive manner. A common problem posed by the application of optogenetic tools is to tune the activity range to be physiologically relevant. Here, we characterized a photoreceptor of the light-oxygen-voltage (LOV) domain family of Phaeodactylum tricornutum aureochrome 1a (AuLOV) as a tool for increasing protein stability under blue light conditions in budding yeast. Structural studies of AuLOVwt, the variants AuLOVM254, and AuLOVW349 revealed alternative dimer association modes for the dark state, which differ from previously reported AuLOV dark-state structures. Rational design of AuLOV-dimer interface mutations resulted in an optimized optogenetic tool that we fused to the photoactivatable adenylyl cyclase from Beggiatoa sp. This synergistic light-regulation approach using two photoreceptors resulted in an optimized, photoactivatable adenylyl cyclase with a cyclic adenosine monophosphate production activity that matches the physiological range of Saccharomyces cerevisiae. Overall, we enlarged the optogenetic toolbox for yeast and demonstrated the importance of fine-tuning the optogenetic tool activity for successful application in cells.

中文翻译：

一种光致工具，用于诱导蛋白质稳定化，以三角角芽孢杆菌Aureochrome 1a轻-氧-电压域为基础。

通过光遗传学工具控制细胞事件是一种以微创方式操纵细胞功能的强大方法。由光遗传学工具的应用引起的一个普遍问题是将活性范围调节为生理学相关的。在这里，我们表征了Phaeodactylum tricornutum aureochrome 1a（AuLOV）的光氧电压（LOV）域家族的光感受器，作为增加蓝芽条件下发芽酵母中蛋白质稳定性的工具。AuLOVwt，变体AuLOVM254和AuLOVW349的结构研究表明，暗态的替代二聚体缔合模式不同于先前报道的AuLOV暗态结构。AuLOV-二聚体界面突变的合理设计导致了优化的光遗传学工具，我们将其融合到了来自Beggiatoa sp。的可光活化的腺苷酸环化酶。使用两个感光器的这种协同光调节方法可产生优化的，可光活化的腺苷酸环化酶，并具有与啤酒酵母的生理范围相匹配的环状单磷酸腺苷生产活性。总体而言，我们扩大了酵母的光遗传学工具箱，并证明了微调光遗传学工具活性对于成功应用于细胞的重要性。

更新日期：2020-02-24

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