Iuliano James N, Gil Agnieszka A, Laptenok Sergey P, Hall Christopher R, Tolentino Collado Jinnette, Lukacs Andras, Hag Ahmed Safaa A, Abyad Jenna, Daryaee Taraneh, Greetham Gregory M, Sazanovich Igor V, Illarionov Boris, Bacher Adelbert, Fischer Markus, Towrie Michael, French Jarrod B, Meech Stephen R, Tonge Peter J
Department of Chemistry, Stony Brook University , Stony Brook, New York 11794, United States.
School of Chemistry, University of East Anglia , Norwich NR4 7TJ, U.K.
Biochemistry. 2018 Feb 6;57(5):620-630. doi: 10.1021/acs.biochem.7b01040. Epub 2018 Jan 4.
The light, oxygen, voltage (LOV) domain proteins are blue light photoreceptors that utilize a noncovalently bound flavin mononucleotide (FMN) cofactor as the chromophore. The modular nature of these proteins has led to their wide adoption in the emerging fields of optogenetics and optobiology, where the LOV domain has been fused to a variety of output domains leading to novel light-controlled applications. In this work, we extend our studies of the subpicosecond to several hundred microsecond transient infrared spectroscopy of the isolated LOV domain AsLOV2 to three full-length photoreceptors in which the LOV domain is fused to an output domain: the LOV-STAS protein, YtvA, the LOV-HTH transcription factor, EL222, and the LOV-histidine kinase, LovK. Despite differences in tertiary structure, the overall pathway leading to cysteine adduct formation from the FMN triplet state is highly conserved, although there are slight variations in rate. However, significant differences are observed in the vibrational spectra and kinetics after adduct formation, which are directly linked to the specific output function of the LOV domain. While the rate of adduct formation varies by only 3.6-fold among the proteins, the subsequent large-scale structural changes in the full-length LOV photoreceptors occur over the micro- to submillisecond time scales and vary by orders of magnitude depending on the different output function of each LOV domain.
光、氧、电压（LOV）结构域蛋白是蓝光光感受器，它们利用非共价结合的黄素单核苷酸（FMN）辅因子作为发色团。这些蛋白的模块化性质使其在光遗传学和光生物学等新兴领域得到广泛应用，在这些领域中，LOV结构域已与多种输出结构域融合，从而产生了新型的光控应用。在这项工作中，我们将对分离的LOV结构域AsLOV2的亚皮秒到数百微秒瞬态红外光谱的研究扩展到三个全长光感受器，其中LOV结构域与一个输出结构域融合：LOV-STAS蛋白YtvA、LOV-HTH转录因子EL222和LOV-组氨酸激酶LovK。尽管三级结构存在差异，但从FMN三重态形成半胱氨酸加合物的总体途径高度保守，尽管速率存在细微差异。然而，在加合物形成后的振动光谱和动力学中观察到了显著差异，这与LOV结构域的特定输出功能直接相关。虽然蛋白质之间加合物形成的速率仅相差3.6倍，但全长LOV光感受器随后的大规模结构变化发生在微秒到亚毫秒的时间尺度上，并且根据每个LOV结构域的不同输出功能相差几个数量级。
