Tertiary Structures of Haseki Tick Virus Nonstructural Proteins Are Similar to Those of Orthoflaviviruses

Abstract

Currently, a large number of novel tick-borne viruses potentially pathogenic to humans are discovered. Studying many of them by classical methods of virology is difficult due to the absence of live viral particles or a sufficient amount of their genetic material. In this case, the use of modern methods of bioinformatics and synthetic and structural biology can help. Haseki tick virus (HSTV) is a recently discovered tick-borne unclassified ssRNA(+) virus. HSTV-positive patients experienced fever and an elevated temperature. However, at the moment, there is no information on the tertiary structure and functions of its proteins. In this work, we used AlphaFold 3 and other bioinformatic tools for the annotation of HSTV nonstructural proteins, based on the principle that the tertiary structure of a protein is inextricably linked with its molecular function. We were the first to obtain models of tertiary structures and describe the putative functions of HSTV nonstructural proteins (NS3 helicase, NS3 protease, NS5 RNA-dependent RNA-polymerase, and NS5 methyltransferase), which play a key role in viral genome replication. Our results may help in further taxonomic identification of HSTV and the development of direct-acting antiviral drugs, POC tests, and vaccines.

Keywords: AlphaFold 3; Flaviviridae; RNA viruses; ixodid ticks; orthoflavi-like viruses; protein structure; tick-borne infection; viral proteins.

Figure 1

Haseki tick virus genome structure and chain topology of the translated single polyprotein. (a) Schematic representation of the complete HSTV genome and (b) nonstructural part of the HSTV genome with chain topology of the translated single polyprotein.

Figure 2

Imposition models of putative NS3 HSTV tertiary structures (ivory) with (a) dengue virus 4, PDB ID: 2VBC (blue), (b) hepatitis C virus, PDB ID: 2F9U (green), and (c) classical swine fever virus, PDB ID: 5WX1 (red).

Figure 3

Putative HSTV NS3 protease tertiary structure: (a) catalytic site (red) and key amino acids of HSTV NS3pro (red circle), (b) hydrophobic clusters (blue) of HSTV NS3pro, (c) electrostatic surface potential of HSTV NS3pro, and (d) imposition models of spatial structure of HSTV NS3pro (ivory) in complex with NSTR1 extracellular domain (red) and Zika virus (PDB ID: 5H6V) NS3pro (grey) in complex with NS2B cofactor (blue). The positive surface potential is colored blue, and the negative surface potential is colored red.

Figure 4

Functional regions of putative HSTV NS3hel. (a) The overall tertiary structure of HSTV NS3hel (ivory) with conservative motifs: motif I (red), motif Ia (green), motif II (blue), motif III (orange), motif IV (cyan), motif IVa (purple), motif V (grey), and motif VI (pink). (b) Sequence alignment of the conservative motifs: red boxes—100% aligned a.a. residues; yellow boxes—80% aligned a.a. residues; white boxes - unaligned. Abbreviations: CSFV—classical swine fever virus; ZIKV—Zika virus; YFV—yellow fever virus; TBEV—tick-borne encephalitis virus; HCV—hepatitis C virus.

Figure 5

Putative NS3 helicase HSTV tertiary structure (ivory): (a) hydrophobic clusters (blue) of HSTV NS3hel, (b) electrostatic surface potential of HSTV NS3hel, and (c) tertiary structure of HSTV NS3hel (ivory) with RNA (red) and ATP. The positive surface potential is colored blue, and the negative surface potential is colored red.

Figure 6

Functional regions of putative HSTV NS5RdRp. (a) The overall tertiary structure of HSTV NS5RdRp (ivory) with catalytic motifs: motif A (blue), motif B (orange), motif C (magenta), motif D (black), motif E (yellow), motif F (green), motif G (red), and priming loop (PL) (cyan). (b) Sequence alignment of the HSTV NS5RdRp motifs: red boxes—100% aligned a.a. residues, yellow boxes—80% aligned a.a. residues, white boxes—unaligned. Abbreviations: CSFV—classical swine fever virus; ZIKV—Zika virus; YFV—yellow fever virus; TBEV—tick-borne encephalitis virus; HCV—hepatitis C virus.

Figure 7

HSTV NS5Mtase structure. (a,b) Imposition models of HSTV NS5MTase spatial structure (ivory) with (a) NS5Mtase Pyrococcus horikoshii, PDB ID: 1WY7 (magenta); (b) NS5Mtase Dengue virus 3, PDB ID: 3P97 (blue); (c) the model of HSTV NS5MTase spatial structure: α-helix (red), β-strand (green); and (d) topology diagram of HSTV NS5MTase: α-helix (red), β-strands (pink), N-amino-terminus, C-carboxyl-terminus.

Figure 8

Spatial structure of putative HSTV NS5Mtase in complex with HSTV NS5-X and HSTV NS5RdRp. (a) Imposition model of HSTV NS5MTase–NS5-X–NS5RdRpspatial structure (gray and ivory) with NS5A zinc-binding domain of hepatitis C virus (green) (PDB ID: 1ZH1) and (b) electrostatic surface potential of HSTV NS5Mtase-NS5-X-NS5RdRp. The positive surface potential is colored blue, and the negative surface potential is colored red.