pH-dependent gating mechanism of the Helicobacter pylori urea channel revealed by cryo-EM.

The urea channel of Helicobacter pylori (HpUreI) is an ideal drug target for preventing gastric cancer but incomplete understanding of its gating mechanism has hampered development of inhibitors for the eradication of H. pylori. Here, we present the cryo-EM structures of HpUreI in closed and open conformations, both at a resolution of 2.7 Å. Our hexameric structures of this small membrane protein (~21 kDa/protomer) resolve its periplasmic loops and carboxyl terminus that close and open the channel, and define a gating mechanism that is pH dependent and requires cooperativity between protomers in the hexamer. Gating is further associated with well-resolved changes in the channel-lining residues that modify the shape and length of the urea pore. Site-specific mutations in the periplasmic domain and urea pore identified key residues important for channel function. Drugs blocking the urea pore based on our structures should lead to a new strategy for H. pylori eradication

Structure of the proton-gated urea channel from the gastric pathogen Helicobacter pylori.

Half the world's population is chronically infected with Helicobacter pylori, causing gastritis, gastric ulcers and an increased incidence of gastric adenocarcinoma. Its proton-gated inner-membrane urea channel, HpUreI, is essential for survival in the acidic environment of the stomach. The channel is closed at neutral pH and opens at acidic pH to allow the rapid access of urea to cytoplasmic urease. Urease produces NH(3) and CO(2), neutralizing entering protons and thus buffering the periplasm to a pH of roughly 6.1 even in gastric juice at a pH below 2.0. Here we report the structure of HpUreI, revealing six protomers assembled in a hexameric ring surrounding a central bilayer plug of ordered lipids. Each protomer encloses a channel formed by a twisted bundle of six transmembrane helices. The bundle defines a previously unobserved fold comprising a two-helix hairpin motif repeated three times around the central axis of the channel, without the inverted repeat of mammalian-type urea transporters. Both the channel and the protomer interface contain residues conserved in the AmiS/UreI superfamily, suggesting the preservation of channel architecture and oligomeric state in this superfamily. Predominantly aromatic or aliphatic side chains line the entire channel and define two consecutive constriction sites in the middle of the channel. Mutation of Trp 153 in the cytoplasmic constriction site to Ala or Phe decreases the selectivity for urea in comparison with thiourea, suggesting that solute interaction with Trp 153 contributes specificity. The previously unobserved hexameric channel structure described here provides a new model for the permeation of urea and other small amide solutes in prokaryotes and archaea

The Ursinus Weekly, November 12, 1917

Student friendship war fund campaign a pronounced success at Ursinus: Students and faculty give generously; goal of $1000 is oversubscribed • Reserves defeat Schuylkill Seminary • Financing the war • Among the colleges • Christian organizations • Literary Societies • On the campushttps://digitalcommons.ursinus.edu/weekly/2538/thumbnail.jp

2007, UMaine News Press Releases

This is a catalog of press releases put out by the University of Maine Division of Marketing and Communications between January 8, 2007 and December 27, 2007

2006, UMaine News Press Releases

This is a catalog of press releases put out by the University of Maine Division of Marketing and Communications between January 4, 2006 and December 19, 2006

New Air-Launched Small Missile (ALSM) Flight Testbed for Hypersonic Systems

A new testbed for hypersonic flight research is proposed. Known as the Phoenix air-launched small missile (ALSM) flight testbed, it was conceived to help address the lack of quick-turnaround and cost-effective hypersonic flight research capabilities. The Phoenix ALSM testbed results from utilization of two unique and very capable flight assets: the United States Navy Phoenix AIM-54 long-range, guided air-to-air missile and the NASA Dryden F-15B testbed airplane. The U.S. Navy retirement of the Phoenix AIM-54 missiles from fleet operation has presented an excellent opportunity for converting this valuable flight asset into a new flight testbed. This cost-effective new platform will fill an existing gap in the test and evaluation of current and future hypersonic systems for flight Mach numbers ranging from 3 to 5. Preliminary studies indicate that the Phoenix missile is a highly capable platform. When launched from a high-performance airplane, the guided Phoenix missile can boost research payloads to low hypersonic Mach numbers, enabling flight research in the supersonic-to-hypersonic transitional flight envelope. Experience gained from developing and operating the Phoenix ALSM testbed will be valuable for the development and operation of future higher-performance ALSM flight testbeds as well as responsive microsatellite small-payload air-launched space boosters

Structure of the master regulator Rns reveals an inhibitor of enterotoxigenic Escherichia coli virulence regulons

Enteric infections caused by the gram-negative bacteria enterotoxigenic Escherichia coli (ETEC), Vibrio cholerae, Shigella flexneri, and Salmonella enterica are among the most common and affect billions of people each year. These bacteria control expression of virulence factors using a network of transcriptional regulators, some of which are modulated by small molecules as has been shown for ToxT, an AraC family member from V. cholerae. In ETEC the expression of many types of adhesive pili is dependent upon the AraC family member Rns. We present here the 3 Å crystal structure of Rns and show it closely resembles ToxT. Rns crystallized as a dimer via an interface similar to that observed in other dimeric AraC’s. Furthermore, the structure of Rns revealed the presence of a ligand, decanoic acid, that inhibits its activity in a manner similar to the fatty acid mediated inhibition observed for ToxT and the S. enterica homologue HilD. Together, these results support our hypothesis that fatty acids regulate virulence controlling AraC family members in a common manner across a number of enteric pathogens. Furthermore, for the first time this work identifies a small molecule capable of inhibiting the ETEC Rns regulon, providing a basis for development of therapeutics against this deadly human pathogen

Transcriptional modulation of enterotoxigenic Escherichia coli virulence genes in response to epithelial cell interactions

Enterotoxigenic Escherichia coli (ETEC) strains are a leading cause of morbidity and mortality due to diarrheal illness in developing countries. There is currently no effective vaccine against these important pathogens. Because genes modulated by pathogen-host interactions potentially encode putative vaccine targets, we investigated changes in gene expression and surface morphology of ETEC upon interaction with intestinal epithelial cells in vitro. Pan-genome microarrays, quantitative reverse transcriptase PCR (qRT-PCR), and transcriptional reporter fusions of selected promoters were used to study changes in ETEC transcriptomes. Flow cytometry, immunofluorescence microscopy, and scanning electron microscopy were used to investigate alterations in surface antigen expression and morphology following pathogen-host interactions. Following host cell contact, genes for motility, adhesion, toxin production, immunodominant peptides, and key regulatory molecules, including cyclic AMP (cAMP) receptor protein (CRP) and c-di-GMP, were substantially modulated. These changes were accompanied by visible changes in both ETEC architecture and the expression of surface antigens, including a novel highly conserved adhesin molecule, EaeH. The studies reported here suggest that pathogen-host interactions are finely orchestrated by ETEC and are characterized by coordinated responses involving the sequential deployment of multiple virulence molecules. Elucidation of the molecular details of these interactions could highlight novel strategies for development of vaccines for these important pathogens

Structure and mechanism of bactericidal mammalian perforin-2, an ancient agent of innate immunity

Perforin-2 (MPEG1) is thought to enable the killing of invading microbes engulfed by macrophages and other phagocytes, forming pores in their membranes. Loss of perforin-2 renders individual phagocytes and whole organisms significantly more susceptible to bacterial pathogens. Here, we reveal the mechanism of perforin-2 activation and activity using atomic structures of pre-pore and pore assemblies, high-speed atomic force microscopy, and functional assays. Perforin-2 forms a pre-pore assembly in which its pore-forming domain points in the opposite direction to its membrane-targeting domain. Acidification then triggers pore formation, via a 180° conformational change. This novel and unexpected mechanism prevents premature bactericidal attack and may have played a key role in the evolution of all perforin family proteins