In 1989, the Hepatitis C virus (HCV) was defined as a non-A, non-B hepatitis virus (Choo et al., 1989). HCV belongs to the Flaviviridae family and genus Hepacivirus, which has a positive single-stranded RNA (Bartenschlager et al., 2011, Moradpour et al., 2007). HCV infection occurs through contact with HCV-infected blood, including blood transfusion, the use of unsterilized medical equipment such as needles, and mother-to-child exposure to infected blood (Guss et al., 2018, Prati, 2006).

In 2005, Wakita et al. created an infectious model based on cultured cells infected with JFH1(a), thereby facilitating numerous subsequent studies on HCV (Wakita et al., 2005). Outside the cells, HCV particles were observed to bind to low-density lipoproteins (LDL) or very low-density lipoproteins (VLDL) and enter cells by binding of the HCV E1 and/or E2 protein with the host cell receptors CD81 and Claudin1. Virus entry is initiated by endocytosis. Upon entering the cell, HCV releases its genomic RNA. The RNA is translated into a polypeptide on the rough endoplasmic reticulum (ER), and the polypeptide is cleaved into the core protein, E1, E2, p7, NS2, NS3, NS4A, NS4B, NS5A, and NS5B with proteases. Core protein, E1 and E2 are defined as structural proteins and are composed of HCV particles, whereas the other proteins are defined as non-structural proteins that facilitate HCV RNA replication. After the HCV genome was translated to polyprotein, signal peptidase mediates the cleavage of C and E1 proteins to generate the whole HCV core. Then HCV core matures upon cleavage with signal peptide peptidase (177 amino acids) (Gawlik and Gallay, 2014). Previous studies have suggested that replicated viral RNA binds to the core protein to create a core-RNA complex and bud into the ER, wearing an envelope including E1 and E2 proteins. Buddled HCV particles are thought to be secreted via the classical protein secretory pathway (Gawlik and Gallay, 2014, Moradpour et al., 2007).

The HCV core protein contains three domains: domains 1, 2, and 3. Domain 1 is a basic amino acid-rich region assumed to bind viral nucleotides (Ivanyi-Nagy et al., 2006, Santolini et al., 1994, Yu et al., 2009). Domain 2 contains amphiphilic alpha helices and contributes to membrane interactions (Boulant et al., 2006). Domain 3 is a transmembrane region cleaved by polyproteins during viral maturation (Pène et al., 2009). The HCV core is thought to play a major role in the HCV life cycle based on its natural binding to RNA. However, it also contributes to RNA binding and interactions with cellular proteins and influences numerous host cell functions (Gawlik and Gallay, 2014, Khaliq et al., 2011). Moriya suggested a relationship between chronic HCV infection and hepatic steatosis and that the viral core protein in frequently observed steatotic changes played a direct role in chronic HCV infection (Moriya et al., 1997). Additionally, interactions with several host cell proteins, which can result in chronic infection and HCV-associated liver pathogenesis, have been reported (Mahmoudvand et al., 2019, Moriishi et al., 2007). However, the structure, RNA binding mechanism, and other biochemical features of HCV remain unclear because an HCV core purification method is lacking, leading to a lack of related structural information.

Boulant et al. (2005), performed structural analysis using a purified HCV core. However, the purification was performed using a denaturant, which could affect the HCV core structure. Therefore, its native structure has not yet been fully elucidated. In this study, we demonstrated purification without denaturation, which provides a more accurate representation of the native structure of the HCV core protein.