Structure of the zinc-binding domain of an essential component of the hepatitis C virus replicase

Abstract

Hepatitis C virus (HCV) is a human pathogen affecting nearly 3% of the world's population¹. Chronic infections can lead to cirrhosis and liver cancer. The RNA replication machine of HCV is a multi-subunit membrane-associated complex. The non-structural protein NS5A is an active component of HCV replicase^2,3, as well as a pivotal regulator of replication^2,4 and a modulator of cellular processes ranging from innate immunity to dysregulated cell growth^5,6. NS5A is a large phosphoprotein (56–58 kDa) with an amphipathic α-helix at its amino terminus that promotes membrane association^7,8,9. After this helix region, NS5A is organized into three domains¹⁰. The N-terminal domain (domain I) coordinates a single zinc atom per protein molecule¹⁰. Mutations disrupting either the membrane anchor^7,8 or zinc binding¹⁰ of NS5A are lethal for RNA replication. However, probing the role of NS5A in replication has been hampered by a lack of structural information about this multifunctional protein. Here we report the structure of NS5A domain I at 2.5-Å resolution, which contains a novel fold, a new zinc-coordination motif and a disulphide bond. We use molecular surface analysis to suggest the location of protein-, RNA- and membrane-interaction sites.