Sandbox Reserved 1769
Sodium-taurocholate Co-transporting Polypeptide
ContentsIntroductionSodium-taurocholate Co-transporting Polypeptide (NTCP) is found within the membrane of liver cells, and its primary role is to facilitate the transport of bile salts into liver cells from the bloodstream. This is important because 90% of human bile salts are recycled daily, so the function of NTCP is critical in providing bile salts to solubilize fats for digestion. In addition to transporting bile salts into the cytoplasm of hepatocytes, NTCP also serves as a receptor for Hepatitis B (HBV) and Hepatitis D (HDV) viruses.
StructureStructures were determined by cryogenic electron microscopy (Cryo-EM) of NTCP in complex with antibodies or nanobodies, revealing two key conformations in NTCP's transport mechanism. There are nine alpha helices spanning the membrane, with the N-terminus located on the extracellular side of the plasma membrane and the C-terminus located on the intracellular side. Transmembrane helices are connected by short loops as well as extracellular and intracellular alpha helices that lie nearly parallel to the membrane. DomainsNTCP contains two characteristic domains: the core and panel domains. Movement of these two domains allows recognition and transport of bile salts into hepatocytes.
Proline/Glycine HingeGlycine and proline residues in the connecting loops and extra- and intracellular helices act as hinges in the mechanism of bile salt uptake. The flexibility allows separation of the core and panel domains, creating a pore open to the extracellular space and exposing critical Na+ binding sites. Once substrate binds the open-pore state, this hinge allows transition to close this pore relative to the extracellular side and open to the cytoplasmic side, thus allowing release of substrate into the cell. Sodium Binding SitesTo transport a single bile salt from the blood to the cytoplasm of the liver cell, two sodium ions are required to be bound to to NTCP in the open-pore state. This is because the transport of bile salts into the cell is so thermodynamically unfavorable that the reaction has to be coupled to the favorable transport of two sodium into into the cell. It is thus an example of secondary active transport (INSERT BLUE LINK). When the bile salts are released into the cell, the protein is then found in the inward facing conformation, in which the pore through which the bile salt had just passed is now closed to the extracellular side. The residues interacting with the sodium ion in sodium binding site #1 include S105, N106, E257, and T123. The residues interacting with the sodium ion in sodium binding site #2 includes Q261 and Q68. Mutations to these significant residues inhibit the binding of sodium ions, and consequently, inhibit the transport of bile salts by NTCP. Significant ResiduesThe majority of residues involved in bile salt uptake are also involved in HBV/HDV infection. Residues 84-87 (extracellular view) of Human NTCP have been shown to be vital for HBV/HDV virus recognition along with bile salt uptake. These residues were replaced in mice NTCP by human NTCP and conferred to successful binding of the virus. These residues are found in the extracellular loop connecting TM2 and TM3. Residues 157-165 (extracellular view) have also been shown to be vital for HBV/HDV viral recognition and bile salt uptake. These residues were mutated in monkey NTCP to the human residues and preS1 binding was then successful. These residues are found on the N-terminal end of TM5. The absence of residues in either of these two extracellular patches hinders preS1 binding and therefore HBV/HDV infection. Interestingly, residues 84-87 do not affect bile acid uptake, so it is a potential site for blocking HBV/HDV infection while maintaining NTCP's ability to perform its normal function. Another important residue was discovered to be a single-nucleotide polymorphism in a small population in East Asia. Residue 267, which is normally serine, being mutated to phenylalanine prevents preS1 binding and does not support bile acid transport. This residue is also found extracellularly, on TM8 of NTCP. There are 3 additional leucine residues that when mutated, block both preS1 binding and HBV/HDV infection. Replacing L27, L31, and L35 (INSERT GREEN LINK) with tryptophan residues presumably blocks the preS1 binding site preventing proper infection. FunctionMechanism of Bile Salt UptakeBile salts recognize and bind to the open-pore state. After binding, bile salts pass through the amphipathic pore (shown below)  and NTCP transitions into the inward facing state. In this conformation, the pore closes off relative to the extracellular side and opens to the cytoplasmic side. Transition to the inward facing state allows release of bile salts and sodium ions. It is not yet known how this transition exactly proceeds. Mechanism of HBV/HDV InfectionHBV and HDV viruses infect are transported through NTCP via secondary active transport. After binding to NTCP in the open-pore state, the viruses remain bound until low bile salt levels in the blood shift equilibria enough that endocytosis of NTCP occurs. Once in the cell, the viruses dissociate and infect. The exact mechanism of how HBV and HDV bind to NTCP is not certain, although two critical sites have been identified on NTCP: residues 84-87 and 157-165. Additionally, it has been shown that myristoylation (INSERT BLUE LINK) of the HBV/HDV capsid is vital for recognition by NTCP, as well as residues 8-17 on HBV/HDV (sequence: NPLGFFPDHQ). (INSERT CITING) has proposed two mechanisms for how HBV/HDV binds to NTCP. The first proposes binding of the myristoyl group to the host cell membrane, while residues 8-17 interact with NTCP residues 157-165. The second proposes binding of the myristoyl group with residues 157-165 in the pore. Medical Relevance
| ||||||||||||
References
Student Contributors
- Ben Minor
- Maggie Samm
- Zac Stanley
