Hepatitis C Virus - Molecular Biology

Molecular Biology

The proteins of this virus are arranged along the genome in the following order: N terminal-core-envelope (E1)-E2-p7-nonstructural protein 2 (NS2)-NS3-NS4A-NS4B-NS5A-NS5B-C terminal. The mature nonstructural proteins (NS2 to NS5B) generation relies on the activity of viral proteinases. The NS2/NS3 junction is cleaved by a metal dependent autocatalytic proteinase encoded within NS2 and the N-terminus of NS3. The remaining cleavages downstream from this site are catalysed by a serine proteinase also contained within the N-terminal region of NS3.

The core protein has 191 amino acids and can be divided into three domains on the basis of hydrophobicity: domain 1 (residues 1-117) contains mainly basic residues with two short hydrophobic regions; domain 2 (resides 118-174) is less basic and more hydrophobic and its C-terminus is at the end of p21; domain 3 (residues 175-191) is highly hydrophobic and acts as a signal sequence for E1 envelope protein.

Both envelope proteins (E1 and E2) are highly glycosylated and important in cell entry. E1 serves as the fusogenic subunit and that E2 acts as the receptor binding protein. E1 has 4-5 N-linked glycans and E2 has 11 N-glycosylation sites.

The p7 protein is dispensable for viral genome replication but plays a critical role in virus morphogenesis. This protein is a 63 amino acid membrane spanning protein which locates itself in the endoplasmic reticulum. Cleavage of p7 is mediated by the endoplasmic reticulum's signal peptidases. Two transmembrane domains of p7 are connected by a cytoplasmic loop and are oriented towards the endoplasmic reticulum's lumen.

NS2 protein is a 21-23 kiloDalton (kDa) transmembrane protein with protease activity.

NS3 is 67 kDa protein whose N-terminal has serine protease activity and whose C-terminal has NTPase/helicase activity. It is located within the endoplasmic reticulum and forms a heterodimeric complex with NS4A - a 54 amino acid membrane protein that acts as a cofactor of the proteinase.

NS4B is a small (27 kDa) hydrophobic integral membrane protein with 4 transmembrane domains. It is located within the endoplasmic reticulum and play an important role for recruitment of other viral proteins. It induces morphological changes to the endoplasmic reticulum forming a structure termed the membranous web.

NS5A is a hydrophilic phosphoprotein which plays an important role in viral replication, modulation of cell signaling pathways and the interferon response. It is known to bind to endoplasmic reticulum anchored human VAP proteins.

The NS5B protein (65 kDa) is the viral RNA dependent RNA polymerase. NS5B has the key function of replicating the HCV’s viral RNA by using the viral positive RNA strand as its template and catalyzes the polymerization of ribonucleoside triphosphates (rNTP) during RNA replication. Several crystal structures of NS5B polymerase in several crystalline forms have been determined based on the same consensus sequence BK (HCV-BK, genotype 1). The structure can be represented by a right hand shape with fingers, palm, and thumb. The encircled active site, unique to NS5B, is contained within the palm structure of the protein. Recent studies on NS5B protein genotype 1b strain J4’s (HC-J4) structure indicate a presence of an active site where possible control of nucleotide binding occurs and initiation of de-novo RNA synthesis. De-novo adds necessary primers for initiation of RNA replication. Current research attempts to bind structures to this active site to alter its functionality in order to prevent further viral RNA replication.

An 11th has also been described. This protein is encoded by a +1 frameshift in the capsid gene. It appears to be antigenic but its function is unknown.