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During HIV-1 virus assembly, the genomic RNA (vRNA) is selected for packaging by the viral protein Gag because it contains a specific packaging signal, Psi. While there have been numerous studies of Gag–Psi interactions, there is almost no information on the kinetic aspects of this interaction. We investigated the kinetics of Gag binding to different RNAs using switchSENSE DRX2 technology. We measured the association rate of Gag binding to monomeric Psi, to a “Multiple Binding Site Mutant” Psi that is inactive for genome packaging in vivo, and to a scrambled Psi. We discovered that Gag binds more rapidly to Psi RNA than to the mutant or scrambled RNAs. Furthermore, rapid Gag association kinetics are retained within sub-regions of Psi: Gag associates more rapidly with RNA containing only the 3′ two of the three Psi stem-loops than with monomeric RNA containing the 5′ two stem-loops or a scrambled RNA. No differences were detectable with individual Psi stem-loops. Interestingly, the rate of binding of Gag molecules to Psi increases with increasing Gag concentration, suggesting cooperativity in binding. The results are consistent with the hypothesis that selectivity in packaging derives from kinetic differences in initiation of particle assembly.

Viral genomic RNA is packaged into virions with high specificity and selectivity. However, in vitro the Gag specificity towards viral RNA is obscured when measured in buffers containing physiological salt. Interestingly, when the binding is challenged by increased salt concentration, the addition of competing RNAs, or introducing mutations to Gag protein, the specificity towards viral RNA becomes detectable. The objective of this work was to examine the contributions of the individual HIV-1 Gag polyprotein domains to nonspecific and specific RNA binding and stability of the initial protein-RNA complexes. Using a panel of Gag proteins with mutations disabling different Gag-Gag or Gag-RNA interfaces, we investigated the distinct contributions of individual domains which distinguish the binding to viral and nonviral RNA by measuring the binding of the proteins to RNAs. We measured the binding affinity in near-physiological salt concentration, and then challenged the binding by increasing the ionic strength to suppress the electrostatic interactions and reveal the contribution of specific Gag–RNA and Gag–Gag interactions. Surprisingly, we observed that Gag dimerization and the highly basic region in the matrix domain contribute significantly to the specificity of viral RNA binding.

The principal structural component of a retrovirus particle is the Gag protein. Retroviral genomic RNAs contain a ‘packaging signal’ (‘Ψ') and are packaged in virus particles with very high selectivity. However, if no genomic RNA is present, Gag assembles into particles containing cellular mRNA molecules. The mechanism by which genomic RNA is normally selected during virus assembly is not understood. We previously reported (Comas-Garcia et al., 2017) that at physiological ionic strength, recombinant HIV-1 Gag binds with similar affinities to RNAs with or without Ψ, and proposed that genomic RNA is selectively packaged because binding to Ψ initiates particle assembly more efficiently than other RNAs. We now present data directly supporting this hypothesis. We also show that one or more short stretches of unpaired G residues are important elements of Ψ; Ψ may not be localized to a single structural element, but is probably distributed over >100 bases.

This article is focused on progressive failure analysis of sandwich structure subjected to transverse low-velocity impact loading. A user defined material model considering non-linear behaviour of the composite skin of the sandwich structure was implemented into commercial software Abaqus by means of material subroutine VUMAT. The experimental measurements of transverse low-velocity impact were performed using an in-house drop tester machine. The results of numerical simulations and experimental data for the sandwich plate deflections at three selected points and the contact force-time dependency during the impact event were compared for particular impact velocities. The occurrence of material damage was visually inspected and compared too.

The contribution describes the methodology of work with human assets, and namely talent management representing a system of work with employees having over-average potential and representing the source of competitive advantage for the company. The work introduces methodological tools for implementing the system of talent management in the following fields: defining the talent, identification of talents, creating the conditions, care at the side of management, individual or group related technical education, development by way of key competences and evaluation of results, both regarding the abilities and skills of individuals through key competences, and by way of indicators of working performance. Describes evaluation using key competences minimal, optimal and actual competence profile and the way of describing competences by behavioural scale. The paper is the part of the research grant MSM 6215648904.