Oligomerization of non-myristoylated M-PMV matrix protein

J. Vlach^1,2, P. Srb^3,1, M. Grocký³, J. Lang^3,1, J. Prchal^1,2, E. Hunter⁴, T. Ruml², R. Hrabal¹

¹Laboratory of NMR Spectroscopy and ²Department of Biochemistry and Microbiology, Institute of Chemical Technology, Prague, Technická 5, 166 28 Prague 6, Czech Republic

³Department of Low Temperature Physics, Faculty of Mathematics and Physics, Charles University, V Holešovičkách 2, 180 00 Prague 8, Czech Republic

⁴Emory Vaccine Center, Yerkes National Primate Research Center, 954 Gatewood Road, Atlanta GA 30329 USA

vlachj@vscht.cz

The self-assembly of viral particle is based on Gag–Gag interactions mediated by homo-interactions of its three domains, i. e. matrix, capsid and nucleocapsid proteins. As the N-terminal part of Gag polyprotein, matrix protein (MA) targets Gag proteins to the site of virus assembly and it directly interacts with plasma membrane, thus initiating budding of viral particles. MAs of most retroviruses are myristoylated on their N termini, which supports the above-mentioned MA functions and strongly promotes MA–MA interactions.

Oligomerization of isolated MA was most extensively studied in Human immunodeficiency virus 1 (HIV-1). It was found that it forms trimers in solution in its myristoylated (myr(+)) rather than non-myristoylated (myr(–)) form [1–3]. Similar oligomerization behaviour was observed in a closely related, though naturally non-myristoylated Equine infectious anaemia virus (EIAV) MA [4].

We studied oligomerization properties of the myr(–) form of Mason-Pfizer monkey virus (M-PMV) matrix protein by means of NMR spectroscopy. Based on dilution-induced chemical shift perturbation data we observed the oligomerization and mapped out the interaction interface of the molecule. A combined quantification of concentration-dependent chemical shifts and translational diffusion coeffcients allowed us to prove for the first time that M-PMV MA exists in a monomer–dimer–trimer equilibrium in solution, and to calculate the respective equilibrium constants. The chemical shift data were further used to calculate structural models of MA oligomers in HADDOCK, revealing a close similarity of M-PMV and HIV-1 MA trimers. The mechanism and possible consequences of M-PMV myr(–)-MA oligomerization will be discussed with respect to the behaviour of HIV-1 and EIAV MAs.

References

1. Y. Morikawa, D. J. Hockley, M. V. Nermut & I. M. Jones, J. Virol., 74 (2000) 16-23.

2. C. Tang, E. Loeliger, P. Luncsford, I. Kinde, D. Beckett & M. F. Summers, Proc. Natl. Acad. Sci. U. S. A., 101 (2004) 517–522.

3. Z. Wu, J. Alexandratos, B. Ericksen, J. Lubkowski, R. C. Gallo & W. Lu, Proc. Natl. Acad. Sci. U. S. A., 101 (2004) 11587–11592.

4. K. Chen, I. Bachtiar, G. Piszczek, F. Bouamr, C. Carter & N. Tjandra, Biochemistry, 47 (2008) 1928–1937.

Acknowledgements

We thank the following institutions for their financial support: the Czech Ministry of Education (Project 1M6837805002), the Grant Agency of Czech Republic (Grant 203/07/0872) and the National Institutes of Health (Grant CA 27834).