Alexey V. Shvetsov

Large-Scale Conformational Flexibility Determines the Properties of AAA+ TIP49 ATPases.

The TIP49a and TIP49b proteins belong to the family of AAA+ ATPases and play essential roles in vital processes such as transcription, DNA repair, snoRNP biogenesis, and chromatin remodeling. We report the crystal structure of a TIP49b hexamer and the comparative analysis of large-scale conformational flexibility of TIP49a, TIP49b, and TIP49a/TIP49b complexes using molecular modeling and molecular dynamics simulations in a water environment. Our results establish key principles of domain mobility that affect protein conformation and biochemical properties, including a mechanistic basis for the downregulation of ATPase activity upon protein hexamerization. These approaches, applied to the lik-TIP49b mutant reported to possess enhanced DNA-independent ATPase activity, help explain how a three-amino acid insertion remotely affects the structure and conformational dynamics of the ATP binding and hydrolysis pocket while uncoupling ATP hydrolysis from DNA binding. This might be similar to the effects of conformations adopted by TIP49 heterohexamers.

Structure of RecX protein complex with the presynaptic RecA filament: Molecular dynamics simulations and small angle neutron scattering

Using molecular modeling techniques we have built the full atomic structure and performed molecular dynamics simulations for the complexes formed by Escherichia coli RecX protein with a single‐stranded oligonucleotide and with RecA presynaptic filament. Based on the modeling and SANS experimental data a sandwich‐like filament structure formed two chains of RecX monomers bound to the opposite sides of the single stranded DNA is proposed for RecX::ssDNA complex. The model for RecX::RecA::ssDNA include RecX binding into the grove of RecA::ssDNA filament that occurs mainly via Coulomb interactions between RecX and ssDNA. Formation of RecX::RecA::ssDNA filaments in solution was confirmed by SANS measurements which were in agreement with the spectra computed from the molecular dynamics simulations.

Method for calculating small-angle neutron scattering spectra using all-atom molecular dynamics trajectories

A method for calculating small-angle neutron scattering (SANS) spectra based on data obtained using the method of the all-atom molecular dynamics of biomacromolecular structures is considered. When interpreting the SANS data, this approach makes it possible to take into account the fact that the structure of biomacromolecules in a solution is not a static object. This method is implemented in the form of a module for the GROMACS software package and will be available in version 4.6 of this popular program package for simulating biomacromolecular-structure dynamics.

The amyloidogenicity of the influenza virus PB1-derived peptide sheds light on its antiviral activity

The influenza virus polymerase complex is a promising target for new antiviral drug development. It is known that, within the influenza virus polymerase complex, the PB1 subunit region from the 1st to the 25th amino acid residues has to be is in an alpha-helical conformation for proper interaction with the PA subunit. We have previously shown that PB1(6-13) peptide at low concentrations is able to interact with the PB1 subunit N-terminal region in a peptide model which shows aggregate formation and antiviral activity in cell cultures. In this paper, it was shown that PB1(6-13) peptide is prone to form the amyloid-like fibrillar aggregates. The peptide homo-oligomerization kinetics were examined, and the affinity and characteristic interaction time of PB1(6-13) peptide monomers and the influenza virus polymerase complex PB1 subunit N-terminal region were evaluated by the SPR and TR-SAXS methods. Based on the data obtained, a hypothesis about the PB1(6-13) peptide mechanism of action was proposed: the peptide in its monomeric form is capable of altering the conformation of the PB1 subunit N-terminal region, causing a change from an alpha helix to a beta structure. This conformational change disrupts PB1 and PA subunit interaction and, by that mechanism, the peptide displays antiviral activity.

Correlated motion of protein subdomains and large-scale conformational flexibility of RecA protein filament

Based on X-ray crystallographic data available at Protein Data Bank, we have built molecular dynamics (MD) models of homologous recombinases RecA from E. coli and D. radiodurans. Functional form of RecA enzyme, which is known to be a long helical filament, was approximated by a trimer, simulated in periodic water box. The MD trajectories were analyzed in terms of large-scale conformational motions that could be detectable by neutron and X-ray scattering techniques. The analysis revealed that large-scale RecA monomer dynamics can be described in terms of relative motions of 7 subdomains. Motion of C-terminal domain was the major contributor to the overall dynamics of protein. Principal component analysis (PCA) of the MD trajectories in the atom coordinate space showed that rotation of C-domain is correlated with the conformational changes in the central domain and N-terminal domain, that forms the monomer-monomer interface. Thus, even though C-terminal domain is relatively far from the interface, its orientation is correlated with large-scale filament conformation. PCA of the trajectories in the main chain dihedral angle coordinate space implicates a co-existence of a several different large-scale conformations of the modeled trimer. In order to clarify the relationship of independent domain orientation with large-scale filament conformation, we have performed analysis of independent domain motion and its implications on the filament geometry.

Triazavirine supramolecular complexes as modifiers of the peptide oligomeric structure

In this study, we present molecular dynamics simulations of the antiviral drug triazavirine, that affects formation of amyloid-like fibrils of the model peptide (SI). According to our simulations, triazavirine is able to form linear supramolecular structures which can act as shields and prevent interactions between SI monomers. This model, as validated by simulations, provides an adequate explanation of triazavirine’s mechanism of action as it pertains to SI peptide fibril formation.

A double-edged sword: supramolecular complexes of triazavirine display multicenter binding effects which influence aggregate formation

Communicated by Ramaswamy H. Sarma

On the structural features of influenza A nucleoprotein particles from small-angle X-ray scattering data

The structure of ribonucleic particles of influenza A virus of the A/California/07/09pdm strain is investigated by transmission electron microscopy and small-angle X-ray scattering. The small-angle X-ray scattering data obtained at room temperature correspond to previously reported data of ribonucleic particles of this virus. At higher temperatures, noticeable changes in the morphology of ribonucleic complexes are observed.

Modeling of conformational transitions of fibrillogenic peptide, homologous to beta-domain of human alpha-lactalbumin

The behavior of the peptide corresponding to beta domain of human alpha-lactalbumin (GYDTQAIVENNESTEYG, WT) has been simulated by the molecular dynamics method. It is shown that, within the model considered, the monomer of this peptide does not tend to form a stable secondary structure; however, simulation of the behavior of several peptide molecules revealed the occurrence of beta structures due to the formation of intermolecular hydrogen bonds. Since the aforementioned interactions involve the terminal portions of peptides, the influence of the tetrapeptide corresponding to the N-terminal portion of WT, TDYG (R), on the secondary structure has been analyzed. The model calculations show that the interaction of this peptide with WT monomer facilitates formation of beta-structures. It is suggested that peptide R may affect the quaternary structure of WT.

Small-angle scattering study of Aspergillus awamori glycoprotein glucoamylase

Glucoamylase from fungus Aspergillus awamori is glycoside hydrolase that catalyzes the hydrolysis of α-1,4- and α-1,6-glucosidic bonds in glucose polymers and oligomers. This glycoprotein consists of a catalytic domain and a starch-binding domain connected by an O-glycosylated polypeptide chain. The conformation of the linker, the relative arrangement of the domains, and the structure of the full-length enzyme are unknown. The structure of the recombinant glucoamylase GA1 was studied by molecular modelling and small-angle neutron scattering (SANS) methods. The experimental SANS data provide evidence that glucoamylase exists as a monomer in solution and contains a glycoside component, which makes a substantial contribution to the scattering. The model of full-length glucoamylase, which was calculated without taking into account the effect of glycosylation, is consistent with the experimental data and has a radius of gyration of 33.4 ± 0.6 Å.