3D models of proteins and conservative G-quadruplexes in the genome of porcine circovirus
Abstract
Background: Non-canonical structures formed in nucleic acid molecules, which include, in particular, multihelical connections 3WJs (three-way junctions), quadruplexes (G4s), are regulatory elements that affect the functioning of the genome. Building 3D models of these structures and pathogen proteins is the first step in understanding their functions. AlphaFold Protein Structure Database was created by Google DeepMind together with EMBL and it contains more than 241 million protein structures. 3D models of porcine circovirus type 3 (PCV-3) proteins have not been identified in this database.
The objective was to identify and characterize potential G4s and 3WJs in the PCV-3 genome, to build 3D
models of PCV-3 proteins.
Materials and Methods: 1138 PCV-3 isolates with complete genomes from GenBank were applied for phylogenetic analysis by MEGA12 software. The BioEdit software was used to construct entropy plots; the QGRS Mapper program was used to search for G4s motifs and determination of their G-scores; the BLAST software was used to search for 100% nucleotide sequence identity. 3D models of proteins and G4s were designed by AlphaFold 3 artificial intelligence.
Results: 4 conserved perfect G4s were identified in the PCV-3 genome, which are formed by three tetrads and confirmed by designing their 3D models. The determined G4s sequences are conservative structural motifs, since the number of PCV-3 isolates in GenBank with G4s in the genome is over 1000. 3D models of G4s the formation of one of which was experimentally determined in the genome of the hepatitis B virus were used as a control of the correctness of these models design. 3D models of the replicase and capsid protein were designed for the two evolutionarily most distant PCV-3 isolates, which were determined from phylogenetic trees based on the cap and rep genes, and the features of their structures were established. 3WJs were not identified in the PCV-3 genome.
Conclusions: 3D models of the replicase, capsid protein of porcine circovirus type 3 as well as four conservative G4s, which are formed by three G-tetrads, were obtained. Increasing the length of the fragments flanking G4s does not prevent their folding, which indicates the importance of these G-rich motifs in the PCV-3 life cycle. The existence of G4s with three tetrads in PCV-3 genome, in contrast to two-tetrad G4s in HBV, in the composition of not only aptamers, but also in molecules of increased length was confirmed by AlphaFold 3.
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