Biophysical analysis of BMV virions purified using a novel method

• Published in: Journal of chromatography. B, Analytical technologies in the biomedical and life sciences Analytical technologies in the biomedical and life sciences, 2017-11, Vol.1068-1069, p.157-163
• Main Authors: Strugała, Aleksander, Kręcisz, Monika, Rybka, Jakub Dalibor, Urbanowicz, Anna, Szpotkowski, Kamil, Bierwagen, Paulina, Figlerowicz, Marek, Kozak, Maciej, Böttcher, Christoph, Giersig, Michał
• Format: Article
• Language: English
• Subjects: Brome mosaic virus; Bromovirus - chemistry; Bromovirus - isolation & purification; Chromatography, Gel; Chromatography, Ion Exchange - methods; Circular Dichroism; Cryogenic transmission electron microscopy; Fourier transform infrared spectroscopy; Hordeum - metabolism; Hordeum - virology; Light; Light scattering; Microscopy, Electron, Transmission; Scattering, Radiation; Spectroscopy, Fourier Transform Infrared; Virion - chemistry; Virion - isolation & purification
• ISSN: 1570-0232; 1873-376X
• DOI: 10.1016/j.jchromb.2017.10.022

•We present a novel purification procedure for BMV.•We confirm high stability of the BMV capsid in pH range from 5.6 to 8.4.•We present results of secondary structure analysis of BMV capsid protein. Brome mosaic virus (BMV) has been successfully loaded with different types of nanoparticles. However, studies concerning its application as a nanoparticle carrier demand high-purity virions in large amounts. Existing BMV purification protocols rely on multiple differential ultracentrifugation runs of the initially purified viral preparation. Herein, we describe an alternative method for BMV purification based on ion-exchange chromatography and size-exclusion chromatography (SEC) yielding 0.2mg of virus from 1g of plant tissue. Our method is of similar efficiency to previously described protocols and can easily be scaled up. The method results in high-quality BMV preparations as confirmed by biophysical analyses, including cryogenic transmission electron microscopy (cryo-TEM), dynamic light scattering (DLS), static light scattering (SLS), and circular dichroism (CD) measurements and attenuated total reflection Fourier transform infrared spectroscopy (ATR-FTIR) spectroscopy. Our results revealed that purified BMV capsids are stable and monodisperse and can be used for further downstream applications. In this work, we also characterize secondary structure and size fluctuations of the BMV virion at different pH values.