Structural and functional characterization of the severe fever with thrombocytopenia syndrome virus L protein

Abstract

The Bunyavirales order contains several emerging viruses with high epidemic potential, including Severe fever with thrombocytopenia syndrome virus (SFTSV). The lack of medical countermeasures, such as vaccines and antivirals, is a limiting factor for the containment of any virus outbreak. To develop such antivirals a profound understanding of the viral replication process is essential. The L protein of bunyaviruses is a multi-functional and multi-domain protein performing both virus transcription and genome replication and, therefore, is an ideal drug target. We established expression and purification procedures for the full-length L protein of SFTSV. By combining single-particle electron cryo-microscopy and X-ray crystallography, we obtained 3D models covering ∼70% of the SFTSV L protein in the apo-conformation including the polymerase core region, the endonuclease and the cap-binding domain. We compared this first L structure of the Phenuiviridae family to the structures of La Crosse peribunyavirus L protein and influenza orthomyxovirus polymerase. Together with a comprehensive biochemical characterization of the distinct functions of SFTSV L protein, this work provides a solid framework for future structural and functional studies of L protein–RNA interactions and the development of antiviral strategies against this group of emerging human pathogens.

Pre-initiation and elongation structures of full-length La Crosse virus polymerase reveal functionally important conformational changes

“…8b). They also diverge from the recently described structures of full-length L protein from Machupo virus (Arenaviridae family) 17 and Severe Fever with Thrombocytopenia Syndrome virus L (Phenuiviridae family) 24,25 published while this manuscript was under review ( Supplementary Fig. 8c, d).…”

The Polarity of an Amino Acid at Position 1891 of Severe Fever with Thrombocytopenia Syndrome Virus L Protein Is Critical for the Polymerase Activity

“…Bunyavirus L protein has two main functional domains: RNA-dependent RNA polymerase activity and endonuclease activity. Previous structural studies reported functional domains of the SFTSV L protein [ 14 , 15 ]. Sequence analysis of the SFTSV YG1 subclones revealed an N1891K amino acid mutation in the L protein located in the C-terminal domain ( Figure 1 A).…”

“…Recent studies revealed the structure of the cap-binding domain of the bunyaviruses including Rift Valley Fever virus (RVFV) and SFTSV, which are located in the C-terminal region of the L protein. This is necessary for cap-snatching alongside the endonuclease domain located at the N-terminal of the L protein [ 14 , 15 , 16 , 26 , 27 ].…”

“…Recent studies have also suggested the presence of potential functional domains in the C-terminal region and conformational structures required for viral transcription initiation, in addition to the cap-binding domain [ 14 , 15 , 16 ]. The results of the minigenome assay are correlated with the electric charge of the of the amino acid sidechain, rather than the size of the sidechain.…”

“…In contrast to studies on the influenza A virus, PA, PB1, and PB2, it remains unclear what functional domains are present in the C-terminal region of the bunyavirus L protein and how they function. Recently, several studies revealed a structural model for the SFTSV L protein, which suggested several domains located at the C-terminal region in addition to the cap-binding domain (CBD) [ 14 , 15 , 16 ].…”