Producing recombinant Zaire ebolavirus Matrix protein VP40 in baculovirus involves co-cloning the gene encoding the full-length VP40 protein (1-326aa) into an expression vector with an N-terminal 10xHis-tag and C-terminal Myc-tag gene, followed by transformation into baculovirus cells. These cells are cultured under conditions that promote protein expression. After sufficient growth is achieved, the cells are lysed to release the recombinant Zaire ebolavirus Matrix protein VP40. Protein purification is performed by affinity chromatography. Its purity is assessed using SDS-PAGE, exceeding 85%.
Zaire ebolavirus Matrix protein VP40 is a crucial component involved in the assembly and budding of the Ebola virus. VP40 plays a central role in virion formation and egress from host cells [1]. It is essential for driving the final stage of Ebola virus replication, which involves the budding of new virus particles from infected cells [2]. VP40 is the most abundant protein in Ebola virus particles, contributing to their filamentous structure and variations in length [3]. The protein facilitates viral budding by interacting with the host cell plasma membrane and promoting the formation of filamentous virus particles [4].
Studies have shown that VP40 forms octamers vital for Ebola virus replication [5]. VP40 has been found to interact with other viral proteins and host factors to mediate specific virus-host interactions necessary for efficient virion release [6]. The protein has also been associated with acetylation modifications that impact its function in virus assembly and release [7].
Furthermore, VP40 has been linked to the organization of the Ebola virus structure, forming a lattice within the viral envelope and interacting with the nucleocapsid to facilitate the assembly of virus particles [8]. It possesses conserved motifs that enable interactions with ubiquitin ligases, further influencing the budding process of filoviruses.
References:
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[3] T. Hoenen, V. Volchkov, L. Kolesnikova, E. Mittler, J. Timmins, M. Ottmannet al., Vp40 octamers are essential for ebola virus replication, Journal of Virology, vol. 79, no. 3, p. 1898-1905, 2005. https://doi.org/10.1128/jvi.79.3.1898-1905.2005
[4] K. Johnson, N. Bhattarai, M. Budicini, C. LaBonia, S. Baker, B. Gerstmanet al., Cysteine mutations in the ebolavirus matrix protein vp40 promote phosphatidylserine binding by increasing the flexibility of a lipid-binding loop, Viruses, vol. 13, no. 7, p. 1375, 2021. https://doi.org/10.3390/v13071375
[5] R. Harty, No exit: targeting the budding process to inhibit filovirus replication, Antiviral Research, vol. 81, no. 3, p. 189-197, 2009. https://doi.org/10.1016/j.antiviral.2008.12.003
[6] D. Hatakeyama, N. Ohmi, A. Saitoh, K. Makiyama, M. Morioka, H. Okazakiet al., Acetylation of lysine residues in the recombinant nucleoprotein and vp40 matrix protein of zaire ebolavirus by eukaryotic histone acetyltransferases, Biochemical and Biophysical Research Communications, vol. 504, no. 4, p. 635-640, 2018. https://doi.org/10.1016/j.bbrc.2018.09.007
[7] D. Beniac, P. Melito, S. deVarennes, S. Hiebert, M. Rabb, L. Lambooet al., The organisation of ebola virus reveals a capacity for extensive, modular polyploidy, Plos One, vol. 7, no. 1, p. e29608, 2012. https://doi.org/10.1371/journal.pone.0029608
[8] R. Harty, M. Brown, G. Wang, J. Huibregtse, & F. Hayes, A ppxy motif within the vp40 protein of ebola virus interacts physically and functionally with a ubiquitin ligase: implications for filovirus budding, Proceedings of the National Academy of Sciences, vol. 97, no. 25, p. 13871-13876, 2000. https://doi.org/10.1073/pnas.250277297
