The Role of Sialyl Glycan Recognition in Host Tissue Tropism of the Avian Parasite Eimeria tenella

Eimeria spp. are a highly successful group of intracellular protozoan parasites that develop within intestinal epithelial cells of poultry, causing coccidiosis. As a result of resistance against anticoccidial drugs and the expense of manufacturing live vaccines, it is necessary to understand the relationship between Eimeria and its host more deeply, with a view to developing recombinant vaccines. Eimeria possesses a family of microneme lectins (MICs) that contain microneme adhesive repeat regions (MARR). We show that the major MARR protein from Eimeria tenella, EtMIC3, is deployed at the parasite-host interface during the early stages of invasion. EtMIC3 consists of seven tandem MAR1-type domains, which possess a high specificity for sialylated glycans as shown by cell-based assays and carbohydrate microarray analyses. The restricted tissue staining pattern observed for EtMIC3 in the chicken caecal epithelium indicates that EtMIC3 contributes to guiding the parasite to the site of invasion in the chicken gut. The microarray analyses also reveal a lack of recognition of glycan sequences terminating in the N-glycolyl form of sialic acid by EtMIC3. Thus the parasite is well adapted to the avian host which lacks N-glycolyl neuraminic acid. We provide new structural insight into the MAR1 family of domains and reveal the atomic resolution basis for the sialic acid-based carbohydrate recognition. Finally, a preliminary chicken immunization trial provides evidence that recombinant EtMIC3 protein and EtMIC3 DNA are effective vaccine candidates

Crystal structure of an orthologue of the NaChBac voltage-gated sodium channel

Voltage-gated sodium (Na(v)) channels are essential for the rapid depolarization of nerve and muscle(1), and are important drug targets(2). A family of bacterial Na(v) channels, exemplified by NaChBac (Na(+)-selective Channel of Bacteria)(3), provides a good model system for structure-function analysis. Here we report the crystal structure of Na(v)AP, a NaChBac orthologue from marine bacteria alpha proteobacterium HIMB114, at 3.05 Å resolution. The channel comprises an asymmetric tetramer. The carbonyl oxygen atoms of Thr178 and Leu179 constitute an inner site within the selectivity filter ((178)TLSSWE(183)) where a Ca(2+) can bind and resides in the crystal structure. The outer mouth of the Na(+) selectivity filter, defined by Ser181 and Glu183, is closed, as is the activation gate at the intracellular side of the pore. The voltage sensors adopt a depolarized conformation with all the gating charges exposing to the extracellular side. We hypothesize that Na(v)AP is captured in an inactivated conformation. Comparison of Na(v)AP with Na(v)Ab(4) reveals significant conformational rearrangements that may underlie the electromechanical coupling mechanism of voltage-gated channels

Specific recognition of RNA/DNA hybrid and enhancement of human RNase H1 activity by HBD

Human RNase H1 contains an N-terminal domain known as dsRHbd for binding both dsRNA and RNA/DNA hybrid. We find that dsRHbd binds preferentially to RNA/DNA hybrids by over 25-fold and rename it as hybrid binding domain (HBD). The crystal structure of HBD complexed with a 12 bp RNA/DNA hybrid reveals that the RNA strand is recognized by a protein loop, which forms hydrogen bonds with the 2′-OH groups. The DNA interface is highly specific and contains polar residues that interact with the phosphate groups and an aromatic patch that appears selective for binding deoxyriboses. HBD is unique relative to non-sequence-specific dsDNA- and dsRNA-binding domains because it does not use positive dipoles of α-helices for nucleic acid binding. Characterization of full-length enzymes with defective HBDs indicates that this domain dramatically enhances both the specific activity and processivity of RNase H1. Similar activity enhancement by small substrate-binding domains linked to the catalytic domain likely occurs in other nucleic acid enzymes

Cryo-EM structure of islet amyloid polypeptide fibrils reveals similarities with amyloid-β fibrils

Amyloid deposits consisting of fibrillar islet amyloid polypeptide (IAPP) in pancreatic islets are associated with beta-cell loss and have been implicated in type 2 diabetes (T2D). Here, we applied cryo-EM to reconstruct densities of three dominant IAPP fibril polymorphs, formed in vitro from synthetic human IAPP. An atomic model of the main polymorph, built from a density map of 4.2-Å resolution, reveals two S-shaped, intertwined protofilaments. The segment 21-NNFGAIL-27, essential for IAPP amyloidogenicity, forms the protofilament interface together with Tyr37 and the amidated C terminus. The S-fold resembles polymorphs of Alzheimer's disease (AD)-associated amyloid-β (Aβ) fibrils, which might account for the epidemiological link between T2D and AD and reports on IAPP-Aβ cross-seeding in vivo. The results structurally link the early-onset T2D IAPP genetic polymorphism (encoding Ser20Gly) with the AD Arctic mutation (Glu22Gly) of Aβ and support the design of inhibitors and imaging probes for IAPP fibrils