Assembly Mechanism of Farnesylated hGBP1 Studied by Time-Resolved Saxs and Electron Microscopy
Assembly Mechanism of Farnesylated hGBP1 Studied by Time-Resolved Saxs and Electron Microscopy
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时间分辨萨克斯和电子显微镜研究法尼基化 hGBP1 的组装机制
DOI：
10.1016/j.bpj.2018.11.880
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发表时间：
2019
影响因子：
3.4
通讯作者：
Andreas M Stadler
中科院分区：
文献类型：
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作者：
Charlotte Lorenz;Andreas M Stadler
蛋白质被优化用于与其他分子和蛋白质的相互作用，从而形成生物体中各种功能的基本基础。自组装如聚集或蛋白质相分离正成为研究焦点，因为故障可能对我们的生活产生严重影响，如阿尔茨海默氏症。我们感兴趣的蛋白质是人鸟苷酸结合蛋白1（hGBP 1），其已知在核苷酸活化后进行同源寡聚化和聚合。在细胞中生理上存在的法尼基脂质的翻译后附着之后，法尼基化的hGBP 1被证明通过在体内形成在没有法尼基修饰的情况下不能观察到的“囊泡样结构”而参与免疫应答，如抗微生物和抗病毒功能[1]。使用时间分辨的超小角X射线散射（TR-USAXS），在核苷酸的存在下的聚合进行了研究，涵盖了广泛的尺寸和时间范围内的光束线ID 02（ESRF）。对于交叉验证，在聚合过程的不同阶段进行时间分辨电子显微镜检查。所获得的成核的真实的空间距离用于小角度散射数据的模型细化。最终模型的散射贡献的单体蛋白质，成核核心，相互作用项和聚合物散射，不同物种的随时间变化的蛋白质聚合的动力学模型中提取和分析。
Proteins are optimized for interactions with other molecules and proteins, thereby forming the fundamental basis for all kind of functions in organisms. Self-assembly as in aggregation or protein phase-separation is becoming a research focus as malfunctioning can have a severe impact on our lives like in the case of Alzheimer's disease. Our protein of interest is the human Guanylate Binding Protein 1 (hGBP1) that is known for homo-oligomerization and polymerization upon nucleotide activation. After post-translational attachment of a farnesyl lipid as physiologically present in cells, the farnesylated hGBP1 was shown to be involved in immune responses like antimicrobial and antiviral functions by forming ‘vesicle-like structures’ in vivo that cannot be observed in absence of the farnesyl modification [1]. Using time-resolved ultra-small angle X-ray scattering (TR-USAXS), the polymerization in presence of nucleotides was studied covering a broad size and time range at the beamline ID02 (ESRF). For cross-validation, time-resolved electron microscopy was performed at different stages of the polymerization process. The obtained real space distances for the nucleation cores were used for a model refinement of the small angle scattering data. With a final model of scattering contributions of monomeric proteins, nucleation cores, an interaction term and polymer scattering, the time-dependent change of the different species are extracted and analyzed in terms of a kinetic model for the protein polymerization.