Structural basis of sodium-dependent bile salt uptake into the liver.

The liver takes up bile salts from blood to generate bile, enabling absorption of lipophilic nutrients and excretion of metabolites and drugs1. Human Na+-taurocholate co-transporting polypeptide (NTCP) is the main bile salt uptake system in liver. NTCP is also the cellular entry receptor of human hepatitis B and D viruses2,3 (HBV/HDV), and has emerged as an important target for antiviral drugs4. However, the molecular mechanisms underlying NTCP transport and viral receptor functions remain incompletely understood. Here we present cryo-electron microscopy structures of human NTCP in complexes with nanobodies, revealing key conformations of its transport cycle. NTCP undergoes a conformational transition opening a wide transmembrane pore that serves as the transport pathway for bile salts, and exposes key determinant residues for HBV/HDV binding to the outside of the cell. A nanobody that stabilizes pore closure and inward-facing states impairs recognition of the HBV/HDV receptor-binding domain preS1, demonstrating binding selectivity of the viruses for open-to-outside over inward-facing conformations of the NTCP transport cycle. These results provide molecular insights into NTCP 'gated-pore' transport and HBV/HDV receptor recognition mechanisms, and are expected to help with development of liver disease therapies targeting NTCP.

Structure of the bile acid transporter and HBV receptor NTCP.

Chronic infection with hepatitis B virus (HBV) affects more than 290 million people worldwide, is a major cause of cirrhosis and hepatocellular carcinoma, and results in an estimated 820,000 deaths annually1,2. For HBV infection to be established, a molecular interaction is required between the large glycoproteins of the virus envelope (known as LHBs) and the host entry receptor sodium taurocholate co-transporting polypeptide (NTCP), a sodium-dependent bile acid transporter from the blood to hepatocytes3. However, the molecular basis for the virus-transporter interaction is poorly understood. Here we report the cryo-electron microscopy structures of human, bovine and rat NTCPs in the apo state, which reveal the presence of a tunnel across the membrane and a possible transport route for the substrate. Moreover, the cryo-electron microscopy structure of human NTCP in the presence of the myristoylated preS1 domain of LHBs, together with mutation and transport assays, suggest a binding mode in which preS1 and the substrate compete for the extracellular opening of the tunnel in NTCP. Our preS1 domain interaction analysis enables a mechanistic interpretation of naturally occurring HBV-insusceptible mutations in human NTCP. Together, our findings provide a structural framework for HBV recognition and a mechanistic understanding of sodium-dependent bile acid translocation by mammalian NTCPs.

Structural insights into the HBV receptor and bile acid transporter NTCP.

Around 250 million people are infected with hepatitis B virus (HBV) worldwide1, and 15 million may also carry the satellite virus hepatitis D virus (HDV), which confers even greater risk of severe liver disease2. The HBV receptor has been identified as sodium taurocholate co-transporting polypeptide (NTCP), which interacts directly with the first 48 amino acid residues of the N-myristoylated N-terminal preS1 domain of the viral large protein3. Despite the pressing need for therapeutic agents to counter HBV, the structure of NTCP remains unsolved. This 349-residue protein is closely related to human apical sodium-dependent bile acid transporter (ASBT), another member of the solute carrier family SLC10. Crystal structures have been reported of similar bile acid transporters from bacteria4,5, and these models are believed to resemble closely both NTCP and ASBT. Here we have used cryo-electron microscopy to solve the structure of NTCP bound to an antibody, clearly showing that the transporter has no equivalent of the first transmembrane helix found in other SLC10 proteins, and that the N terminus is exposed on the extracellular face. Comparison of our structure with those of related proteins indicates a common mechanism of bile acid transport, but the NTCP structure displays an additional pocket formed by residues that are known to interact with preS1, presenting new opportunities for structure-based drug design.