Evidence for alternative quaternary structure in a bacterial Type III secretion system chaperone

<p>Abstract</p> <p>Background</p> <p>Type III secretion systems are a common virulence mechanism in many Gram-negative bacterial pathogens. These systems use a nanomachine resembling a molecular needle and syringe to provide an energized conduit for the translocation of effector proteins from the bacterial cytoplasm to the host cell cytoplasm for the benefit of the pathogen. Prior to translocation specialized chaperones maintain proper effector protein conformation. The class II chaperone, Invasion plasmid gene (Ipg) C, stabilizes two pore forming translocator proteins. IpgC exists as a functional dimer to facilitate the mutually exclusive binding of both translocators.</p> <p>Results</p> <p>In this study, we present the 3.3 Å crystal structure of an amino-terminally truncated form (residues 10-155, denoted IpgC^10-155) of the class II chaperone IpgC from <it>Shigella flexneri</it>. Our structure demonstrates an alternative quaternary arrangement to that previously described for a carboxy-terminally truncated variant of IpgC (IpgC^1-151). Specifically, we observe a rotationally-symmetric "head-to- head" dimerization interface that is far more similar to that previously described for SycD from <it>Yersinia enterocolitica </it>than to IpgC^1-151. The IpgC structure presented here displays major differences in the amino terminal region, where extended coil-like structures are seen, as opposed to the short, ordered alpha helices and asymmetric dimerization interface seen within IpgC^1-151. Despite these differences, however, both modes of dimerization support chaperone activity, as judged by a copurification assay with a recombinant form of the translocator protein, IpaB.</p> <p>Conclusions</p> <p>From primary to quaternary structure, these results presented here suggest that a symmetric dimerization interface is conserved across bacterial class II chaperones. In light of previous data which have described the structure and function of asymmetric dimerization, our results raise the possibility that class II chaperones may transition between asymmetric and symmetric dimers in response to changes in either biochemical modifications (e.g. proteolytic cleavage) or other biological cues. Such transitions may contribute to the broad range of protein-protein interactions and functions attributed to class II chaperones.</p

Johnson Space Center's regenerative life support systems test bed

The Regenerative Life Support System (RLSS) Test Bed at NASA's Johnson Space Center is an atmospherically closed, controlled environment facility for the evaluation of regenerative life support systems using higher plants in conjunction with physicochemical life support systems. When completed, the facility will be comprised of two large scale plant growth chambers, each with approximately 10 m(exp 2) growing area. One of the two chambers, the Variable Pressure Growth Chamber (VPGC), will be capable of operating at lower atmospheric pressures to evaluate a range of environments that may be used in Lunar or Martian habitats; the other chamber, the Ambient Pressure Growth Chamber (APGC) will operate at ambient atmospheric pressure. The root zone in each chamber will be configurable for hydroponic or solid state media systems. Research will focus on: (1) in situ resource utilization for CELSS systems, in which simulated lunar soils will be used in selected crop growth studies; (2) integration of biological and physicochemical air and water revitalization systems; (3) effect of atmospheric pressure on system performance; and (4) monitoring and control strategies

Signaling Virtue or Vulnerability? The Changing Impact of Exchange Rate Regimes on Government Bond Yields

Do exchange rate regimes affect the conditions under which developed countries borrow? This paper argues that they do, but their impact on yields depends on the prevailing macroeconomic context. When investors regard inflation as the most relevant risk to bond holdings, monetary union has a distinct advantage over floating and fixed exchange rates because of its credible in-built mechanism to control inflation. However, once default is seen as the most relevant risk, exchange rate rigidity becomes a liability due to its constraining effect on governments’ ability to respond to adverse shocks. We test our argument with a moving window panel analysis for twenty-three OECD countries from 1980 to 2017. We find that before the late 2000s, inflation was penalized under floating and (to a lesser extent) fixed exchange rate regimes, but not in countries in monetary union. Since the 2010s, inflation carries no penalty under any exchange rate regime. Variables linked to default risk (debt and entitlement spending) did not affect yields under any exchange rate arrangements until the mid-2000s. Afterwards, countries in monetary union (and to a lesser extent in fixed exchange rate regimes) were significantly penalized for public debt and entitlement spending, whereas countries with floating regimes were not. Our results speak to the literatures on governments’ institutional commitments and “room to move.”Haben Wechselkursregime einen Einfluss auf die Konditionen, zu denen entwickelte Länder Staatsanleihen ausgeben können? Wir argumentieren in diesem Beitrag, dass dies der Fall ist, wobei ihre Wirkung auf die Anleiherenditen vom vorherrschenden makroökonomischen Kontext abhängt. Erachten Investoren Inflation als das entscheidende Risiko für Investitionen in Anleihen, so hat eine Währungsunion durch ihren glaubwürdigen inte­grierten Mechanismus zur Inflationskontrolle klare Vorteile gegenüber flexiblen und festen Wechselkursen. Wird jedoch ein Ausfall der Rückzahlungen als das entscheidende Risiko angesehen, werden starre Wechselkurse zum Nachteil, da sie die Fähigkeit von Regierungen, auf negative Schocks zu reagieren, verringern. Wir testen unser Argument mithilfe einer für den Zeitraum von 1980 bis 2017 mit gleitenden Zeitfenstern durchgeführten Panelana­lyse von 23 OECD-Ländern. Die Ergebnisse zeigen, dass Inflation vor den späten 2000er-Jahren in flexiblen und (weniger stark) in festen Wechselkursregimen finanziell abgestraft wurde, jedoch nicht in den Ländern einer Währungsunion. Seit den 2010er-Jahren wirkt sich Inflation in keinem der Wechselkursregime auf die Renditen aus. Mit dem Ausfallri­siko verknüpfte Variablen (Staatsverschuldung und Sozialausgaben) hatten bis zur Mitte der 2000er-Jahre in keinem der Wechselkursregime einen Einfluss auf die Renditen. Danach wurden Länder in einer Währungsunion erheblich (und Länder in festen Wechselkursregimen weniger stark) für Staatsverschuldung und Sozialausgaben abgestraft, während dies bei Ländern in flexiblen Regimen nicht der Fall war. Unsere Ergebnisse tragen zur Literatur über institutionelle Selbstverpflichtungen und Handlungsspielräume von Regierungen bei.Contents 1 Introduction 2 Commitment devices and policy autonomy from a theoretical perspective 3 Inflation risk, default risk, and exchange rate regimes: An analysis of twenty-three OECD countries 4 Results section Results Robustness checks 5 Discussion and conclusion Appendix A Sources used for identifying sample countries’ exchange rate regimes over time Appendix B Results controlling for capital mobility (measured via the Chinn-Ito capital account openness index) Appendix C Results excluding countries with a greater share of foreign currency-denominated debt than 2 percent (Canada and Sweden) Appendix D Results for crawling peg exchange rate regime Appendix E Results excluding heavily indebted EMU countries (Belgium, Greece, and Italy) Appendix F Results excluding EMU countries with the lowest sovereign credit ratings prior to the crisis (Greece, Italy, and Portugal) Reference

Computational modeling of TC0583 as a putative component of the Chlamydia muridarum V-type ATP synthase complex and assessment of its protective capabilities as a vaccine antigen.

Numerous Chlamydia trachomatis proteins have been identified as potential subunit vaccines, of which the major outer-membrane protein (MOMP) has, so far, proven the most efficacious. Recently, subunit A of the V-type ATP synthase (ATPase; TC0582) complex was shown to elicit partial protection against infection. Computational modeling of a neighboring gene revealed a novel subunit of the V-type ATPase (TC0583). To determine if this newly identified subunit could induce protection and/or enhance the partial protection provided by subunit A alone, challenge studies were performed using a combination of these recombinant proteins. The TC0583 subunit alone and concurrently with TC0582, was used to vaccinate BALB/c mice utilizing CpG-1826 and Montanide ISA 720 VG as adjuvants. Vaccinated animals were challenged intranasally with Chlamydia muridarum and the course of the infection was followed. Mice immunized with individual antigens showed minimal alleviation of body weight reduction; however, mice immunized with TC0583 and TC0582 in combination, displayed weight loss levels close to those observed with MOMP. Importantly, immunization with a combination of recombinant subunit proteins reduced chlamydial inclusion forming units by approximately a log-fold. These protection levels support that, these highly conserved Chlamydia proteins, in combination with other antigens, may serve as potential vaccine candidates

Direct Catalytic N-Alkylation of α-Amino Acid Esters and Amides Using Alcohols with High Retention of Stereochemistry

The direct functionalization of naturally abundant chiral scaffolds such as α-amino acid esters or amides with widely abundant alcohols, without any racemization, is a demanding transformation that is of central importance for the synthesis of bio-active compounds. Herein a robust and general method was developed for the direct N-alkylation of α-amino acid esters and amides with alcohols. This powerful ruthenium-catalyzed methodology is atom-economic, base-free, and allowed for excellent retention of stereochemical integrity. The use of diphenylphosphate as additive was crucial for significantly enhancing reactivity and product selectivity. Notably, the only by-product was water and both substrates could be potentially derived from renewable resources

Major Histocompatibility Complex Class I- And II-Deficient Knock-Out Mice Are Resistant to Primary but Susceptible to Secondary Eimeria Papillata Infections

Two distinct mechanisms seem to function in reducing oocyst output during Eimeria papillata infections in mice. For naive mice, immunity was afforded by a T-cell-independent gamma-interferon (IFN-γ) response mediated by natural killer (NK) cells. On reinfection, resistance was associated with T-cells and, to a lesser extent, perforin. To determine if antigen presentation with major histocompatibility complex (MHC) molecules was required to control oocyst production by NK cells during primary infection or by T-cells during secondary infection, mutant mice that lacked H2-IAβb (Aβb(-/-)) or β2-microglobulin (β2m(-/-)) were used. Since MHC molecules are required for the maturation of αβ T-cells, Aβb(-/-) and β2m(-/-) mutant mice are also deficient in functional αβ+ CD4+ or αβ+ CD8+ T-cells, respectively. As compared with wild-type control mice, oocyst output by mutant mice was not significantly affected during primary infection, suggesting that the ability of NK cells to control parasite replication is not dependent on the expression of MHC molecules. On reinfection, differences were observed for mutant mice as compared with controls. Aβb(-/-) mice were found to be more susceptible than β2m(-/-) mice, suggesting that the αβ+ CD4+ T-cell subset plays a greater role in resistance to reinfection than does the αβ+ CD8+ T-cell subset. The mechanism of resistance depends on the immune status of the host and requires the coordinated interaction of both αβ+ T-cell, subsets for optimal parasite control during subsequent infections

Performance of Sweetpotato for Bioregenerative Life Support

Sweetpotato was successfully grown to harvest maturity in a large-scale atmospherically-closed controlled environment chamber. Yield of edible biomass and capacity for contributing to air revitalization and water recovery were documented. Yield was slightly less than that found in smaller-scale studies, but this is not unusual (Wheeler 1999). Continued work is suggested to improve control of storage root initiation, bulking and vine growth

Orally Active MMP-1 Sparing α-tetrahydropyranyl and α-piperidinyl Sulfone Matrix Metalloproteinase (MMP) Inhibitors with Efficacy in Cancer, Arthritis, and Cardiovascular Disease

α-Sulfone-α-piperidine and α-tetrahydropyranyl hydroxamates were explored that are potent inhibitors of MMP’s-2, -9, and -13 that spare MMP-1, with oral efficacy in inhibiting tumor growth in mice and left-ventricular hypertrophy in rats and in the bovine cartilage degradation ex vivo explant system. α-Piperidine 19v (SC-78080/SD-2590) was selected for development toward the initial indication of cancer, while α-piperidine and α-tetrahydropyranyl hydroxamates 19w (SC-77964) and 9i (SC-77774), respectively, were identified as backup compounds

Structural and functional studies of type three secretion virulence factors from gram-negative pathogenic bacteria

Title from PDF of title page, viewed on December 21, 2011Dissertation advisor: Brian GeisbrechtVitaIncludes bibliographic references (p. 93-98)Thesis (Ph.D.)--School of Biological Sciences. University of Missouri-Kansas City, 2011Many pathogenic Gram-negative bacteria utilize type III secretion systems (TTSS) to alter the normal functions of target host epithelial cells. Of the 1.1 million deaths that are caused by Shigella each year, nearly a third are in children under five years of age. Salmonella enterica serovar Typhimurium is the leading cause of hospitalization and death due to food-borne gastroenteritis in the U.S. The pathogenesis of both of these species involves the invasion of epithelial cells of the gastrointestinal tract, which requires the use of a type III secretion system (TTSS). The Shigella type III secretion apparatus (TTSA) is composed of a basal body spanning both bacterial membranes and an exposed oligomeric needle. Host altering effectors are secreted through this energized conduit to promote bacterial invasion. The active needle-tip complex of S. flexneri is composed of a tip protein, IpaD, and two pore-forming translocators, IpaB and IpaC. Maturation of the needle-tip complex proceeds in a stepwise manner. IpaD is at the tip of the nascent TTSA needle where it controls the first step of TTS activation. The bile salt deoxycholate (DOC) binds to IpaD to induce recruitment of the first translocator, IpaB, into the maturing tip complex. The pore-forming translocators are bound by the class II chaperone, IpgC, within the bacterial cytoplasm in order to prevent premature association and degradation. Despite their importance in promoting Shigella virulence, few molecular level details are known regarding the interactions between IpgC and its targets, IpaB and IpaC. Additionally, the mechanism by which DOC serves to stabilize a conformational change within IpaD is poorly understood. Methods in structural biology, in particular X-ray crystallography, are extremely valuable in addressing such questions. We present here the crystal structures of IpgC (identifying an alternative quaternary state), DOC-bound IpaD and the N-terminal regions of both IpaB and SipB, the S. Typhimurium first translocator homolog. These structures have facilitated the functional analysis of crucial determinants of Gram-negative pathogens that would otherwise not have been possible. Additionally, these structural studies have revealed the critical α-helical nature of each protein subunit involved in mature needle-tip-translocon formation.Introduction -- Materials and methodology -- Evidence for alternative quaternary structure in a bacterial type III secretion system chaperone -- Identification of the bile salt binding site on Ipad from shigella flexneri and the influence of ligand binding on Ipad structure -- The crystal structures of coiled-coil domains from type three secretion system first translocator proteins reveal homology to pore-forming toxins -- Global discussio