Fakhouri Hussein, Mas Caroline, Le Roy Aline, Marchal Estelle, Pasquier Coralie, Diat Olivier, Bauduin Pierre, Ebel Christine
ICSM, CEA, CNRS, ENSCM, Univ Montpellier, UMR 5257 - Bâtiment 426 - Centre de Marcoule BP 17171, BAGNOLS SUR CEZE CEDEX, 30207, Marcoule, France.
Université Grenoble Alpes, CNRS, CEA, EMBL, ISBG, 38000, Grenoble, France.
Eur Biophys J. 2025 Apr 23. doi: 10.1007/s00249-025-01746-y.
The self-assembly of the cobaltabis(dicarbollide) (COSAN) anionic boron clusters into micelles above a critical micelle concentration (cmc) of 10-20 mM and its behavior as "sticky nano-ions" facilitating controlled protein aggregation have been previously investigated using scattering techniques. These techniques effectively provide average structural parameters but, when applied to colloidal systems, often rely on models assuming polydispersity or anisotropic shapes. Here, we employed sedimentation velocity analytical ultracentrifugation (SV-AUC), which offers the ability to resolve discrete species. We revisited two key questions: (1) the aggregation behavior of COSAN into micelles, a topic still under debate, and (2) the nature of the protein assemblies induced by COSAN, specifically their size/shape distribution and aggregation number. SV-AUC confirms the cmc of COSAN of 16 mM and reveals that COSAN micelles exhibit low aggregation numbers (8 in water and 14 in dilute salt), consistent with recent hypotheses. It shows that COSAN promotes myoglobin aggregation into discrete oligomeric species with well-defined aggregation numbers, such as dimers, tetramers, and higher-order assemblies, depending on the COSAN-to-protein ratio. COSAN binding could be quantified at the lower COSAN/myoglobin ratios. For example, at ratio 5, myoglobin monomer (25%) binds about two COSANs, dimer (45%) about 14 COSANs, and there are ≈ 30% very large aggregates. These results provide clarity on the discrete nature of COSAN micelle aggregation and protein assembly. This study highlights the complementary role of SV-AUC in understanding supramolecular assemblies, offering useful insights into the behavior of COSAN nano-ions and their interactions with biomacromolecules.
先前已使用散射技术研究了钴双（二碳硼烷）（COSAN）阴离子硼簇在临界胶束浓度（cmc）为10 - 20 mM以上时自组装成胶束的过程及其作为“粘性纳米离子”促进可控蛋白质聚集的行为。这些技术有效地提供了平均结构参数，但应用于胶体系统时，通常依赖于假设多分散性或各向异性形状的模型。在此，我们采用沉降速度分析超速离心法（SV - AUC），它能够分辨离散的物种。我们重新审视了两个关键问题：（1）COSAN聚集成胶束的聚集行为，这一话题仍在争论中；（2）COSAN诱导的蛋白质聚集体的性质，特别是它们的尺寸/形状分布和聚集数。SV - AUC证实了COSAN的cmc为16 mM，并揭示COSAN胶束的聚集数较低（在水中为8，在稀盐中为14），这与最近的假设一致。结果表明，COSAN促进肌红蛋白聚集形成具有明确聚集数的离散寡聚体物种，如二聚体、四聚体和高阶聚集体，这取决于COSAN与蛋白质的比例。在较低的COSAN/肌红蛋白比例下可以对COSAN结合进行定量。例如，在比例为5时，肌红蛋白单体（25%）结合约两个COSAN，二聚体（45%）结合约14个COSAN，并且存在约30%的非常大的聚集体。这些结果为COSAN胶束聚集和蛋白质组装的离散性质提供了清晰的认识。本研究突出了SV - AUC在理解超分子组装中的互补作用，为COSAN纳米离子的行为及其与生物大分子的相互作用提供了有用的见解。
