Oligonucleotides antiviral drugs have been actively implemented in medicine during the last decades nevertheless the molecular mechanism of their action is still unclear. As it was shown in our previous work, the combination of oligonucleotides with alcohol sugar D-mannitol leads to changes in their biological activity and efficiency. At this stage of our investigation, we studied the ability of oligoribonucleotides from total yeast RNA (ORNs) and oligoribonucleotides-D-mannitol complexes to affect the conformation and stability of interferon (IFN) α-2b – a key protein of the antiviral cell defense mechanism.
To investigate that interaction, conformational changes and stability of IFN protein in the presence and/or absence of the ligands were studied by fluorescence and CD (circular dichroism) spectroscopies. All experiments were performed on spectrofluorometer Jasco FP- 8200 and CD spectrometer Jasco J-815 with a Peltier temperature cell holder. Obtained thermal denaturation profiles of IFN α–2b alone and in the presence of ORNs and ORNs-D-mannitol complexes show that the addition of these ligands led to an increase of thermal stabilization of protein of 2 and 1.8 0C respectively. The dissociation constant between INF and total yeast ORNs was Kd =2.88±1.14µM and between INF and ORNs with D-mannitol – Kd =0,92±0.23µM.
The analysis of IFN secondary structure changes by Bestsel shows that addition of ORNs or ORNs-D-mannitol complexes led to a decrease of α-helix components in the protein structure and to an increase of antiparallel β-stand, β-turn, and random coil components. At the same time, the analysis of the tertiary structure shows that adding ORNs-D-mannitol changes the architecture of the protein from the 2-layer sandwich to alpha-beta complex. On the other hand, adding ORNs did not cause any change in the tertiary structure.
We suppose that total yeast ORNs and ORNs-D-mannitol complexes act as compounds, altering the secondary and tertiary structures of the interferon and in this way can change its biological activity.
