RESUMO
Three 'triple gene block' proteins known as TGBp1, TGBp2 and TGBp3 are required for cell-to-cell movement of plant viruses belonging to a number of genera including Hordeivirus. Hordeiviral TGBp1 interacts with viral genomic RNAs to form ribonucleoprotein (RNP) complexes competent for translocation between cells through plasmodesmata and over long distances via the phloem. Binding of hordeivirus TGBp1 to RNA involves two protein regions, the C-terminal NTPase/helicase domain and the N-terminal extension region. This study demonstrated that the extension region of hordeivirus TGBp1 consists of two structurally and functionally distinct domains called the N-terminal domain (NTD) and the internal domain (ID). In agreement with secondary structure predictions, analysis of circular dichroism spectra of the isolated NTD and ID demonstrated that the NTD represents a natively unfolded protein domain, whereas the ID has a pronounced secondary structure. Both the NTD and ID were able to bind ssRNA non-specifically. However, whilst the NTD interacted with ssRNA non-cooperatively, the ID bound ssRNA in a cooperative manner. Additionally, both domains bound dsRNA. The NTD and ID formed low-molecular-mass oligomers, whereas the ID also gave rise to high-molecular-mass complexes. The isolated ID was able to interact with both the NTD and the C-terminal NTPase/helicase domain in solution. These data demonstrate that the hordeivirus TGBp1 has three RNA-binding domains and that interaction between these structural units can provide a basis for remodelling of viral RNP complexes at different steps of cell-to-cell and long-distance transport of virus infection.