Aggregation of Influenza A Virus Nuclear Export Protein

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A. O. Golovko^1,a, O. N. Koroleva^2,b*, A. P. Tolstova^3,c, N. V. Kuz’mina^4,5,d, E. V. Dubrovin^3,e, and V. L. Drutsa^6,f

¹Lomonosov Moscow State University, Department of Bioengineering and Bioinformatics, 119991 Moscow, Russia

²Lomonosov Moscow State University, Department of Chemistry, 119991 Moscow, Russia

³Lomonosov Moscow State University, Department of Physics, 119991 Moscow, Russia

⁴Lomonosov Moscow State University, Department of Biology, 119991 Moscow, Russia

⁵Frumkin Institute of Physical Chemistry and Electrochemistry, Russian Academy of Sciences, 119071 Moscow, Russia

⁶Lomonosov Moscow State University, Belozersky Research Institute of Physico-Chemical Biology, 119991 Moscow, Russia

^* To whom correspondence should be addressed.

Received April 12, 2018; Revision received July 4, 2018

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Influenza A virus nuclear export protein (NEP) plays an important role in the viral life cycle. Recombinant NEP proteins containing (His)₆-tag at either N- or C-terminus were obtained by heterologous expression in Escherichia coli cells and their high propensity for aggregation was demonstrated. Dynamic light scattering technique was used to study the kinetics and properties of NEP aggregation in solutions under different conditions (pH, ionic strength, presence of low-molecular-weight additives and organic solvents). Using atomic force microscopy, the predominance of spherical aggregates in all examined NEP preparations was shown, with some amyloid-like structures being observed in the case of NEP-C protein. A number of structure prediction programs were used to identify aggregation-prone regions in the NEP structure. All-atom molecular dynamics simulations indicate a high rate of NEP molecule aggregation and reveal the regions preferentially involved in the intermolecular contacts that are located at the edges of the rod-like protein molecule. Our results suggest that NEP aggregation is determined by different types of interactions and represents an intrinsic property of the protein that appears to be necessary for its functioning in vivo.

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KEY WORDS: influenza A virus, nuclear export protein NEP, protein aggregation

DOI: 10.1134/S0006297918110111

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