Defining the Stoichiometry and Cargo Load of Viral and Bacterial Nanoparticles by Orbitrap Mass Spectrometry

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• 作者：Joost Snijder ; Michiel van de Waterbeemd ; Eugen Damoc ; Eduard Denisov ; Dmitry Grinfeld ; Antonette Bennett ; Mavis Agbandje-McKenna ; Alexander Makarov ; Albert J. R. Heck
• 刊名：Journal of the American Chemical Society
• 出版年：2014
• 出版时间：May 21, 2014
• 年：2014
• 卷：136
• 期：20
• 页码：7295-7299
• 全文大小：320K
• 年卷期：v.136,no.20(May 21, 2014)
• ISSN：1520-5126

文摘

Accurate mass analysis can provide useful information on the stoichiometry and composition of protein-based particles, such as virus-like assemblies. For applications in nanotechnology and medicine, such nanoparticles are loaded with foreign cargos, making accurate mass information essential to define the cargo load. Here, we describe modifications to an Orbitrap mass spectrometer that enable high mass analysis of several virus-like nanoparticles up to 4.5 MDa in mass. This allows the accurate determination of the composition of virus-like particles. The modified instrument is utilized to determine the cargo load of bacterial encapsulin nanoparticles that were engineered to encapsulate foreign cargo proteins. We find that encapsulin packages from 8 up to 12 cargo proteins, thereby quantifying cargo load but also showing the ensemble spread. In addition, we determined the previously unknown stoichiometry of the three different splice variants of the capsid protein in adeno-associated virus (AAV) capsids, showing that symmetry is broken and assembly is heterogeneous and stochastic. These results demonstrate the potential of high-resolution mass analysis of protein-based nanoparticles, with widespread applications in chemical biology and nanotechnology.