Loic Delcourte, Corinne Sanchez, Estelle Morvan, Mélanie Berbon, Axelle Grélard, Claire Saragaglia, Thierry Dakhli, Stéphane Thore, Benjamin Bardiaux, Birgit Habenstein, Brice Kauffmann, Sven J. Saupe, Antoine Loquet
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引用次数: 0
摘要
信号体是一种高阶蛋白质机制,参与控制调节免疫防御和细胞死亡执行的复杂机制。免疫反应是通过识别外源或内源信号,触发信号体组装过程而启动的。信号体命运的最后一步往往涉及膜靶向和激活孔形成执行域,导致膜破坏,最终导致细胞死亡。在植物、哺乳动物和真菌中,这种诱导细胞死亡的结构域已被彻底表征,特别是真菌细胞死亡执行蛋白结构域 HeLo。然而,人们对细菌中基于信号体的免疫反应机制知之甚少,而且对细菌信号体中细胞死亡执行蛋白的构象仍然知之甚少。我们最近发现了多种多细胞细菌中存在的 NLR 信号体,并利用基因组挖掘方法鉴定了橄榄色链霉菌中的推定细胞死亡执行体。这些蛋白质包含一个参与信号传递的 C 端淀粉样结构域和一个 N 端结构域(BELL,表示细菌类似于真菌的 HeLL(HeLo-like)),可能负责膜靶向、孔形成和细胞死亡执行。在本研究中,我们报告了 S. olivochromogenes BELL2 的高产率表达及其溶液核磁共振光谱特性。BELL 在溶液中折叠,我们报告了骨架和侧链的分配。我们发现了五个α螺旋二级结构元素和一个折叠核心,其体积远小于其真菌同源物 HeLo。这项研究为全长蛋白质的组装及其膜靶向的核磁共振研究迈出了第一步。
NMR resonance assignment of the cell death execution domain BELL2 from multicellular bacterial signalosomes
Signalosomes are high-order protein machineries involved in complex mechanisms controlling regulated immune defense and cell death execution. The immune response is initiated by the recognition of exogeneous or endogenous signals, triggering the signalosome assembly process. The final step of signalosome fate often involves membrane-targeting and activation of pore-forming execution domains, leading to membrane disruption and ultimately cell death. Such cell death-inducing domains have been thoroughly characterized in plants, mammals and fungi, notably for the fungal cell death execution protein domain HeLo. However, little is known on the mechanisms of signalosome-based immune response in bacteria, and the conformation of cell death executors in bacterial signalosomes is still poorly characterized. We recently uncovered the existence of NLR signalosomes in various multicellular bacteria and used genome mining approaches to identify putative cell death executors in Streptomyces olivochromogenes. These proteins contain a C-terminal amyloid domain involved in signal transmission and a N-terminal domain, termed BELL for Bacteria analogous to fungal HeLL (HeLo-like), presumably responsible for membrane-targeting, pore-forming and cell death execution. In the present study, we report the high yield expression of S. olivochromogenes BELL2 and its characterization by solution NMR spectroscopy. BELL is folded in solution and we report backbone and sidechain assignments. We identified five α-helical secondary structure elements and a folded core much smaller than its fungal homolog HeLo. This study constitutes the first step toward the NMR investigation of the full-length protein assembly and its membrane targeting.
期刊介绍:
Biomolecular NMR Assignments provides a forum for publishing sequence-specific resonance assignments for proteins and nucleic acids as Assignment Notes. Chemical shifts for NMR-active nuclei in macromolecules contain detailed information on molecular conformation and properties.
Publication of resonance assignments in Biomolecular NMR Assignments ensures that these data are deposited into a public database at BioMagResBank (BMRB; http://www.bmrb.wisc.edu/), where they are available to other researchers. Coverage includes proteins and nucleic acids; Assignment Notes are processed for rapid online publication and are published in biannual online editions in June and December.