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.
Extra-cytoplasmic function (ECF) σ-factors are widespread in bacteria, linking environmental stimuli with changes in gene expression. These transcription factors span several phylogenetically distinct groups and are remarkably diverse in their activation and regulatory mechanisms. Here, we describe the structural and biochemical features of a Mycobacterium tuberculosis ECF factor σJ that suggests that the SnoaL_2 domain at the C-terminus can modulate the activity of this initiation factor in the absence of a cognate regulatory anti-σ factor. M. tuberculosis σJ can bind promoter DNA in vitro; this interaction is substantially impaired by the removal of the SnoaL_2 domain. This finding is consistent with assays to evaluate σJ-mediated gene expression. Structural similarity of the SnoaL_2 domain with epoxide hydrolases also suggests a novel functional role for this domain. The conserved sequence features between M. tuberculosis σJ and other members of the ECF41 family of σ-factors suggest that the regulatory mechanism involving the C-terminal SnoaL_2 domain is likely to be retained in this family of proteins. These studies suggest that the ECF41 family of σ-factors incorporate features of both—the σ70 family and bacterial one—component systems thereby providing a direct mechanism to implement environment-mediated transcription changes.
Mycobacterium tuberculosis has multiple σ factors which enable the bacterium to reprogram its transcriptional machinery under diverse environmental conditions. σ(J), an extracytoplasmic function σ factor, is upregulated in late stationary phase cultures and during human macrophage infection. σ(J) governs the cellular response to hydrogen peroxide-mediated oxidative stress. σ(J) differs from other canonical σ factors owing to the presence of a SnoaL_2 domain at the C-terminus. σ(J) crystals belonged to the tetragonal space group I422, with unit-cell parameters a = b = 133.85, c = 75.08 Å. Diffraction data were collected to 2.16 Å resolution on the BM14 beamline at the European Synchrotron Radiation Facility (ESRF).
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