Abstract Zika virus (ZIKV) Non-structural protein 1 (NS1) plays an essential role in viral replication and immune evasion. Our understanding of the differential protective mechanism of NS1-targeting antibodies is limited. Here, we determined the cryoEM structures of ZIKV NS1 in complex with two group antibodies at 2.6-2.9Å. Group I antibodies (3G2 and 4B8) potently recognize cell surface form of NS1 and multiple oligomeric forms of NS1 by occupy the epitopes on outer surface of dimeric NS1. IgG and Fab from group I antibodies completely abrogate sNS1-mediated endothelial dysfunction in vitro. Group II antibodies (4F10, 2E11, and 14G5) recognize the previously reported epitopes in distal end of the β -ladder domain of monomeric NS1, and their blockade efficiency depends on the affinity with NS1 protein. These findings elucidate the correlation between the epitope recognition and the protective efficacy of anti-NS1 antibodies and highlight the distinct mechanisms of therapeutic potential of 3G2 and 4B8.
Yellow fever virus (YFV) infections can cause severe diseases in humans, resulting in mass casualties in Africa and the Americas each year. Secretory NS1 (sNS1) is thought to be used as a diagnostic marker of flavivirus infections, playing an essential role in the flavivirus life cycle, but little is known about the composition and structure of YFV sNS1. Here, we present that the recombinant YFV sNS1 exists in a heterogeneous mixture of oligomerizations, predominantly in the tetrameric form. The cryoEM structures show that the YFV tetramer of sNS1 is stacked by the hydrophobic interaction between β-roll domains and greasy fingers. According to the 3D variability analysis, the tetramer is in a semi-stable state that may contain multiple conformations with dynamic changes. We believe that our study provides critical insights into the oligomerization of NS1 and will aid the development of NS1-based diagnoses and therapies.
Abstract Dihydropyridine receptor (DHPR), an L-type Ca 2+ channel complex, plays an essential role in muscle contraction, secretion, integration of synaptic input in neurons and synaptic transmission. The molecular architecture of DHPR complex remains elusive. Here we present a 15-Å resolution cryo-electron microscopy structure of the skeletal DHPR/L-type Ca 2+ channel complex. The DHPR has an asymmetrical main body joined by a hook-like extension. The main body is composed of a “trapezoid” and a “tetrahedroid”. Homologous crystal structure docking and site-specific antibody labelling revealed that the α1 and α2 subunits are located in the “trapezoid” and the β subunit is located in the “tetrahedroid”. This structure revealed the molecular architecture of a eukaryotic Ca 2+ channel complex. Furthermore, this structure provides structural insights into the key elements of DHPR involved in physical coupling with the RyR/Ca 2+ release channel and shed light onto the mechanism of excitation-contraction coupling.
Abstract The glucagon receptor family comprises Class B G protein-coupled receptors (GPCRs) that play a crucial role in regulating blood sugar levels. Receptors of this family represent important therapeutic targets for the treatment of diabetes and obesity. Despite intensive structural studies, we only have a poor understanding of the mechanism of peptide hormone-induced Class B receptor activation. This process involves the formation of a sharp kink in transmembrane helix 6 that moves out to allow formation of the nucleotide-free G protein complex. Here, we present the cryo-EM structure of the glucagon receptor (GCGR), a prototypical Class B GPCR, in complex with an engineered soluble glucagon derivative and the heterotrimeric G-protein, G s . Comparison with the previously determined crystal structures of GCGR bound to a partial agonist reveals a structural framework to explain the molecular basis of ligand efficacy that is further supported by mutagenesis data.
Abstract The type-1 ryanodine receptor (RyR1) is an intracellular calcium release channel for skeletal muscle excitation-contraction coupling. Published structures of RyR1 showed RyR1 is open only in the assistance of exogenous regulators, such as caffeine, ryanodine, PCB-95 and diamide. Here, we report that with a mild purification procedure, a single transmembrane protein junctin is co-purified with RyR1. RyR-junctin complex can be activated to open state by Ca2+ only. Junctin inserts its transmembrane helix next to S1, S4 and S4-5 linker of RyR1, exerting an out-ward pushing force to the TM domain, facilitating the dilation of S6. Junctin regulates the channel conformation by directly affecting the TM domain via Jun/S1-4–S4-5 linker. Additional ATP increases the channel open probability by further stabilizing the channel in the open state. Our results demonstrate junctin forms an intrinsic complex with RyR1 and play a key role in the channel gating of RyR1 in physiological context.
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