Neorhodopsins, Near-infrared sensing Rhodopsin-cyclases

新视紫红质、近红外传感视紫红质环化酶

• 批准号: 509731234
• 负责人: Professor Dr. Franz Bartl
• 金额: --
• 依托单位: Institut für Biologie
• 依托单位国家: 德国
• 项目类别: Research Grants
• 资助国家: 德国
• 项目状态: 未结题
• 来源: https://gepris.dfg.de/gepris/projekt/509731234?language=en
• 关键词: Neorhodopsins; Near; infrared; sensing; Rhodopsin

Microbial rhodopsins are key proteins in optogenetics, since theyenable the control of cellular activity by external light. Until now, optogenetic experiments or resulting therapies in living organismshave been limited because the activating light is strongly absorbed by the surrounding tissue. The use of near-infrared (NIR) light is a potential solution for this problem, but so far rhodopsins were not sensitive for this absorption region. Neorhodopsin (NeoR), a novel fungal Rhodopsin-cyclase that we recently discovered is capable of absorbing NIR light for the first time and therefore serves as a template for opening this spectral range for other rhodopsins. In addition to its unusual NIR absorption, NeoR has the unique features of being highly fluorescent and forming functional heterodimers. Besides NeoR, these rhodopsin-cyclase complexes contain a conventional rhodopsin that absorbs blue or green light and thereby activates the cyclase. Homodimeric Rhodopsin-cyclases with sensitivity in the green spectral region have been previouslydescribed, whereas heterodimeric rhodopsin complexes are hitherto completely unknown. NeoR is a bistable photoreceptor and is photoswitchable between a NIR and a UV absorbing state, thus affecting the enzyme activity in a yet unknown way. The aim of the proposed project is to elucidate the functioning of NeoR in the heterodimeric photoreceptor addressing the following questions: What are the structural features that cause these novel Rhodopsin-cyclases to function only as heterodimers? How does the light-induced state of the NeoR subunit affect the cyclase activity? In addition, we aim to study the photochemistry of NeoR to understand how it differs from conventional rhodopsins. For these studies, we are using different experimental approaches, such as the kinetic characterization of the enzyme reaction, as well as the elucidation of light-induced structural changes using static and time-resolved spectroscopy. Furthermore, we want to investigate homologous proteins from related fungi in order to identify heterodimeric Rhodopsin-cyclase suitable for optogenetic applications and to find NeoR variants with further red-shifted absorption. To understand the physiological role of these heterodimeric photoreceptors and in particular of NeoR, we intend to study the photo-orientation of motile fungal zoospores, in analogy to function of homodimeric Rhodopsin-cyclase. Detailed knowledge of the structure and function of the NIR-active chromophore in NeoR will enable strategies for making this spectral region accessible to optogenetics, which would allow completely new applications.

微生物视紫红质是光遗传学中的关键蛋白质，因为它们可以通过外部光控制细胞活动。到目前为止，光遗传学实验或由此产生的生物体疗法受到限制，因为激活光被周围组织强烈吸收。使用近红外（NIR）光是这个问题的潜在解决方案，但到目前为止，视紫红质对该吸收区域不敏感。新视紫红质（NeoR）是我们最近发现的一种新型真菌视紫红质环化酶，它首次能够吸收近红外光，因此可以作为其他视紫红质打开该光谱范围的模板。除了其不寻常的近红外吸收之外，NeoR 还具有高荧光和形成功能性异二聚体的独特功能。除了 NeoR 之外，这些视紫红质-环化酶复合物还含有传统的视紫红质，可以吸收蓝光或绿光，从而激活环化酶。先前已经描述了在绿色光谱区域具有敏感性的同二聚视紫红质环化酶，而异二聚视紫红质复合物迄今为止是完全未知的。 NeoR 是一种双稳态光感受器，可在近红外和紫外吸收状态之间进行光切换，从而以未知的方式影响酶活性。该项目的目的是阐明 NeoR 在异二聚体光感受器中的功能，解决以下问题：导致这些新型视紫红质环化酶仅作为异二聚体发挥作用的结构特征是什么？ NeoR 亚基的光诱导状态如何影响环化酶活性？此外，我们的目标是研究 NeoR 的光化学，以了解它与传统视紫红质的不同之处。在这些研究中，我们使用不同的实验方法，例如酶反应的动力学表征，以及使用静态和时间分辨光谱阐明光诱导的结构变化。此外，我们希望研究来自相关真菌的同源蛋白，以确定适合光遗传学应用的异二聚体视紫红质环化酶，并找到具有进一步红移吸收的 NeoR 变体。为了了解这些异二聚光感受器，特别是 NeoR 的生理作用，我们打算研究运动真菌游动孢子的光定向，类似于同二聚视紫红质环化酶的功能。对 NeoR 中近红外活性发色团的结构和功能的详细了解将有助于制定使该光谱区域可用于光遗传学的策略，从而实现全新的应用。