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Structure-function analysis of coxsackie B3 virus protein 2B
F J Van kuppeveld1, W J Melchers, K Kirkegaard
1Department of Medical Microbiology, University of Nijmegen, The Netherlands.
Abstract:
Expression of poliovirus protein 2B in mammalian cells inhibits protein secretion and increases the susceptibility of the cells to hygromycin B, consistent with the increase in plasma membrane permeability seen during poliovirus infection (J. R. Doedens and K. Kirkegaard, EMBO J. 14, 894-907, 1995). We report here that expression of protein 2B of the closely related coxsackie B3 virus (CBV3) leads to the same biochemical alterations. Analysis of several mutant CBV3 2B proteins that contain mutations in a predicted cationic amphipathic alpha-helix (F. J. M. van Kuppeveld, J. M. D. Galama, J. Zoll, P. J. J. C. van den Hurk, and W. J. G. Melchers, J. Virol. 70, 3876-3886, 1996) demonstrated that the integrity of this domain is crucial for both biochemical functions of 2B. Mutations in a second hydrophobic domain (F. J. M. van Kuppeveld, J. M. D. Galama, J. Zoll, and W. J. G. Melchers, J. Virol. 69, 7782-7790, 1995), on the other hand, are more disruptive to the ability of CBV3 2B to inhibit protein secretion than to increase membrane permeability. Therefore, inhibition of protein secretion is not merely a consequence of the membrane changes that increase uptake of hygromycin B. The existence of mutations that interfere with virus growth but do not impair the ability of 2B to inhibit protein secretion or increase membrane permeability argues for additional functions of protein 2B.
Insights
Coxsackievirus B3 protein 2B alters mammalian cells by inhibiting protein secretion and increasing membrane permeability. Specific domains within protein 2B are crucial for these functions, suggesting additional roles beyond membrane changes.
Area of Science:
- Virology
- Molecular Biology
- Cell Biology
Background:
- Poliovirus protein 2B inhibits protein secretion and increases cell membrane permeability.
- These effects are observed in mammalian cells and are linked to poliovirus infection.
Purpose of the Study:
- To investigate if coxsackievirus B3 (CBV3) protein 2B induces similar biochemical alterations in mammalian cells.
- To determine the functional importance of specific domains within CBV3 protein 2B for its biochemical activities.
Main Methods:
- Expression of wild-type and mutant CBV3 protein 2B in mammalian cells.
- Analysis of protein secretion inhibition and hygromycin B susceptibility.
- Mutagenesis of predicted cationic amphipathic alpha-helix and hydrophobic domains within CBV3 2B.
Main Results:
- CBV3 protein 2B expression inhibits protein secretion and increases membrane permeability, mirroring poliovirus 2B effects.
- The cationic amphipathic alpha-helix domain is essential for both functions.
- Mutations in a second hydrophobic domain differentially affect protein secretion inhibition versus membrane permeability.
Conclusions:
- Inhibition of protein secretion by CBV3 2B is not solely a consequence of increased membrane permeability.
- CBV3 protein 2B likely possesses functions beyond altering membrane permeability and inhibiting protein secretion.
- Specific domains within CBV3 2B are critical for its distinct biochemical activities.