Structural and functional characterization of the CAP domain of pathogen-related yeast 1 (Pry1) protein

The production, crystal structure, and functional characterization of the C-terminal cysteine-rich secretory protein/antigen 5/pathogenesis related-1 (CAP) domain of pathogen-related yeast protein-1 (Pry1) from Saccharomyces cerevisiae is presented. The CAP domain of Pry1 (Pry1CAP) is functional in vivo as its expression restores cholesterol export to yeast mutants lacking endogenous Pry1 and Pry2. Recombinant Pry1CAP forms dimers in solution, is sufficient for in vitro cholesterol binding, and has comparable binding properties as full-length Pry1. Two crystal structures of Pry1CAP are reported, one with Mg2+ coordinated to the conserved CAP tetrad (His208, Glu215, Glu233 and His250) in spacegroup I41 and the other without divalent cations in spacegroup P6122. The latter structure contains four 1,4-dioxane molecules from the crystallization solution, one of which sits in the cholesterol binding site. Both structures reveal that the divalent cation and cholesterol binding sites are connected upon dimerization, providing a structural basis for the observed Mg2+-dependent sterol binding by Pry1

Schistosoma mansoni venom allergen-like protein 4 (SmVAL4) is a novel lipid-binding SCP/TAPS protein that lacks the prototypical CAP motifs

Schistosomiasis is a parasitic disease that affects over 200 million people. Vaccine candidates have been identified, including Schistosoma mansoni venom allergen- like proteins (SmVALs) from the SCP/TAPS (sperm-coating protein/Tpx/antigen 5/pathogenesis related-1/Sc7) superfamily. The first SmVAL structure, SmVAL4, was refined to a resolution limit of 2.16 Å. SmVAL4 has a unique structure that could not be predicted from homologous structures, with longer loops and an unusual C-terminal extension. SmVAL4 has the characteristic ***Missing image substitution***/***Missing image substitution***-sandwich and central SCP/TAPS cavity. Furthermore, SmVAL4 has only one of the signature CAP cavity tetrad amino-acid residues and is missing the histidines that coordinate divalent cations such as Zn²⁺ in other SCP/TAPS proteins. SmVAL4 has a cavity between ***Missing image substitution***-helices 1 and 4 that was observed to bind lipids in tablysin-15, suggesting the ability to bind lipids. Subsequently, SmVAL4 was shown to bind cholesterol in vitro. Additionally, SmVAL4 was shown to complement the in vivo sterol-export phenotype of yeast mutants lacking their endogenous CAP proteins. Expression of SmVAL4 in yeast cells lacking endogenous CAP function restores the block in sterol export. These studies suggest an evolutionarily conserved lipid-binding function shared by CAP proteins such as SmVAL4 and yeast CAP proteins such as Pry1

Crystal structure of MpPR-1i, a SCP/TAPS protein from Moniliophthora perniciosa , the fungus that causes witches’ broom disease of cacao

The pathogenic fungi Moniliophthora perniciosa causes Witches’ Broom Disease (WBD) of cacao. The structure of MpPR-1i, a protein expressed by M. perniciosa when it infects cacao, are presented. This is the first reported de novo structure determined by single-wavelength anomalous dispersion phasing upon soaking with selenourea. Each monomer has flexible loop regions linking the core alpha-beta-alpha sandwich topology that comprise ~50% of the structure, making it difficult to generate an accurate homology model of the protein. MpPR-1i is monomeric in solution but is packed as a high ~70% solvent content, crystallographic heptamer. The greatest conformational flexibility between monomers is found in loops exposed to the solvent channel that connect the two longest strands. MpPR-1i lacks the conserved CAP tetrad and is incapable of binding divalent cations. MpPR-1i has the ability to bind lipids, which may have roles in its infection of cacao. These lipids likely bind in the palmitate binding cavity as observed in tablysin-15, since MpPR-1i binds palmitate with comparable affinity as tablysin-15. Further studies are required to clarify the possible roles and underlying mechanisms of neutral lipid binding, as well as their effects on the pathogenesis of M. perniciosa so as to develop new interventions for WBD

Crystal Structure of Borrelia turicatae protein, BTA121, a differentially regulated gene in the tick-mammalian transmission cycle of relapsing fever spirochetes

Tick-borne relapsing fever (RF) borreliosis is a neglected disease that is often misdiagnosed. RF species circulating in the United States include Borrelia turicatae, which is transmitted by argasid ticks. Environmental adaptation by RF Borrelia is poorly understood, however our previous studies indicated differential regulation of B. turicatae genes localized on the 150 kb linear megaplasmid during the tick- mammalian transmission cycle, including bta121. This gene is up-regulated by B. turicatae in the tick versus the mammal, and the encoded protein (BTA121) is predicted to be surface localized. The structure of BTA121 was solved by single- wavelength anomalous dispersion (SAD) using selenomethionine-derivative protein. The topology of BTA121 is unique with four helical domains organized into two helical bundles. Due to the sequence similarity of several genes on the megaplasmid, BTA121 can serve as a model for their tertiary structures. BTA121 has large interconnected tunnels and cavities that can accommodate ligands, notably long parallel helices, which have a large hydrophobic central pocket. Preliminary in-vitro studies suggest that BTA121 binds lipids, notably palmitate with a similar order of binding affinity as tablysin-15, a known palmitate-binding protein. The reported data will guide mechanistic studies to determine the role of BTA121 in the tick-mammalian transmission cycle of B. turicatae

DETEKSI GLUKOSA DALAM URIN ORANGUTAN SUMATERA (PONGO ABELII) MENGGUNAKAN STRIPTEST SEMIKUANTITATIF DI TAMAN HEWAN PEMATANG SIANTAR

Penelitian ini bertujuan mengetahui ada tidaknya glukosa dalam uron orangutan sumatera (Pongo abelii) sebagai penunjang diagnosa di Taman Hewan Pematang Siantar, Sumatera Utara. Pengoleksian urin terhadap 4 ekor orangutan sumatera di dalam kandang yang dilakukan pada pagi hari yaitu saat orangutan bangun tidur atau sebelum pemberian pakan orangutan. Pengulangan uji dilakukan 3 kali selama 10 hari pada bulan Januari 2015. Setelah pengoleksian urin kemudian dilakukan pemeriksaan dengan cara mencelupkan stripstest pada 5-10 ml urin selama 30 detik. Analisis data menggunakan metode deskriptif kualitatif dengan hasil bersifat semikuantitatif melalui pembacaan nilai glukosa pada stripstest yang memiliki skala perubahan warna yaitu : negatif, positif 1 (100 mg/dL), positif 2 (250 mg/dL), positif 3 (500 mg/dL), dan positif 4 (1000 mg/dL). Hasil penelitian menunjukkan bahwa dari 4 sampel urin orangutan sumatera tidak terdeteksi adanya glukosa dalam urin

A People-Centred Social Totality Approach to Low-Income Housing in the Developing World

In a significant part of the developing world, especially sub-Saharan Africa, public housing policies and implementation have depended on a top-to-bottom approach in an attempt to ensure housing supply. However, public authorities sometimes backed by international agencies preferring to operate through the housing market have failed to meet the housing need, especially for low-income people. Even when the users are involved like in the slum dwellers association, the organisation of the process is majorly controlled by the public authorities. While government and public institutions attained minimal success in housing provision for the lowest classes in the society, the people have been more successful in housing production. This chapter situates the housing problem and policy responses in the context of the developing world characterised by limited capacity to control and manage the largely more successful informal people-controlled housing production structure. A cyclic people-centred strategy framework for low-income housing is proposed based on town-gown collaboration in studying low-income people, their activated housing process and the houses produced to guide present strategies and synthesise future strategies and policy. This framework emanates from Henri Lefebvre’s social totality explanation to understand how low-income people negotiate housing from the social context

A Culture-Based Design Pedagogy for Nigerian and South African Spatial Forms

Previous authors have discussed non-Western design forms with regards to the significance of integrating global issues, diversity issues in design curricula, and designing in diverse cultural settings. However, there are few studies that examine instructional approaches that use non-Western African design forms. The purpose of this study was to develop and test instruction on Nigerian and South African spatial forms in an Interior Design studio in a Southwestern University (N=17). The study focused on how students responded to the instruction, their ability to synthesize design ideas for different cultural settings using design theories, their utilization of examples from non-Western perspectives as references for discussing design, and their ability to solve design problems in a different cultural setting. The hope was that the extent to which their skills improved will significantly prepare them for a diverse and global society.The instruction was developed using one facet of ACT-R learning theory (Anderson, 1995), anthropological methods (Creswell, 2009; Hall, 1966; Kingsolver, 1998; O'Reilly, 2005; Silverman, 2005), and Grant's pedagogical approaches (Grant, 1991). The distinction between declarative and procedural knowledge was used to help students learn about Nigerian and South African spatial forms and how to apply those forms. Anthropological methods were used to elucidate information about Nigerian and South African design aesthetics. Grant's (1991) pedagogical approach of introducing diversity in design education was embedded in the instruction using three steps: the inclusion, contribution, and transformational approaches.An ethnographic study which combined a case study methodology from educational research and the comparative method from anthropology encapsulated the experiences of participants. The data sources were pre- and post-test questionnaires, observational data, video recording, actual design projects developed by participants, and interview data. Multiple data sources indicated the instructional design process was successful in helping students problem-solve in a different cultural setting. It highlighted the importance of helping students with the development of declarative knowledge on Nigeria and South Africa, teacher-centered and discovery methods, and constant feedback as a way to foster automatization (Anderson, 1995).Participants responded positively to the instruction. The data showed they used a combination of abstract and concrete themes derived from Nigerian and South African cultures to develop their design solutions. Participants demonstrated their understanding of diverse background of design theories in their creative thinking, critical thinking and decision-making processes during the study. This was evident in how the different student groups articulated their spatial organization, implemented aspects from the cultures artistic expressions in their solutions, and demonstrated an understanding of color and materials from the different cultures. Evidence from different data sources such as the questionnaires, observational data, and interviews show that students were able to use examples from non-Western perspectives as references for discussing design ideas. Additionally, through multiple data sources, students report being better at solving design problems in a different cultural setting

X-ray structures of Na-GST-1 and Na-GST-2 two glutathione s-transferase from the human hookworm Necator americanus

<p>Abstract</p> <p>Background</p> <p>Human hookworm infection is a major cause of anemia and malnutrition of adults and children in the developing world. As part of on-going efforts to control hookworm infection, The Human Hookworm Vaccine Initiative has identified candidate vaccine antigens from the infective L3 larval stages and adult stages of the parasite. Adult stage antigens include the cytosolic glutathione-S-transferases (GSTs). Nematode GSTs facilitate the inactivation and degradation of a variety of electrophilic substrates (drugs) via the nucleophilic addition of reduced glutathione. Parasite GSTs also play significant roles in multi-drug resistance and the modulation of host-immune defense mechanisms.</p> <p>Results</p> <p>The crystal structures of <it>Na</it>-GST-1 and <it>Na</it>-GST-2, two major GSTs from <it>Necator americanus </it>the main human hookworm parasite, have been solved at the resolution limits of 2.4 Å and 1.9 Å respectively. The structure of <it>Na</it>-GST-1 was refined to R-factor 18.9% (R-free 28.3%) while that of <it>Na</it>-GST-2 was refined to R-factor 17.1% (R-free 21.7%). Glutathione usurped during the fermentation process in bound in the glutathione binding site (G-site) of each monomer of <it>Na</it>-GST-2. <it>Na</it>-GST-1 is uncomplexed and its G-site is abrogated by Gln 50. These first structures of human hookworm parasite GSTs could aid the design of novel hookworm drugs.</p> <p>Conclusion</p> <p>The 3-dimensional structures of <it>Na</it>-GST-1 and <it>Na</it>-GST-2 show two views of human hookworm GSTs. While the GST-complex structure of <it>Na</it>-GST-2 reveals a typical GST G-site that of <it>Na</it>-GST-1 suggests that there is some conformational flexibility required in order to bind the substrate GST. In addition, the overall binding cavities for both are larger, more open, as well as more accessible to diverse ligands than those of GSTs from organisms that have other major detoxifying mechanisms. The results from this study could aid in the design of novel drugs and vaccine antigens.</p

Intestinal schistosomiasis in an apparently healthy rural population in Bayelsa State, Nigeria

Background: Schistosomiasis is endemic in Nigeria and three species; Schistosoma haematobium, Schistosoma mansoni, and Schistosoma intercalatum have been reported in Niger Delta, Nigeria. This study aimed to determine the prevalence of schistosomiasis in rural communities of Bayelsa State, Nigeria.Methodology: Four rural homogeneous communities; Otuegala, Immiringi, Otuesega, and Ibelebiri in Ogbia Local Government Area of Bayelsa State, Nigeria, were randomly selected for the study. A structured questionnaire was administered to each participant in their native language and used to collect participant’s biodata and swimming history. Stool samples collected from all participants were examined qualitatively by wet preparation and after formolethol concentration. Data were analyzed using SPSS version 20.0 software and results presented in proportion and tables.Results: A total of 829 participants (age range 1 - 80 years) were recruited for the study. Helminth ova were identified in the stool samples of 218 (26.3%) participants. Among 380 males examined, 82 (21.6%) were infected, while out of 449 females examined, 138 (30.3%) were infected. The ova of seven helminths identified and their frequency of occurrence were; S. intercalatum 86 (10.4%), Ascaris lumbricoides 53 (6.4%), S. mansoni 35 (4.2%), Trichuris trichiura 22 (2.6%), hookworm 20 (2.4%) and Taenia spp 2 (0.2%). Schistosoma haematobium was identified in non-urine contaminated stool sample of an eight-year old boy. A total of 11 (1.3%) participants had double infections, affecting 7 (63.6%) females and 4 (36.4%) males, with the commonest combination being S. intercalatum and A.lumbricoides 6 (0.7%), followed by S. intercalatum and hookworm 4 (0.5%), and S. mansoni and hookworm 1(0.1%).Conclusion: S. intercalatum was the most prevalent intestinal helminthic infection in this study, which is a rare finding in most epidemiological investigations. The affinity of Schistosoma species to establish double infections with hookworm and other intestinal helminths should be taken into account during chemoprophylaxis. Keywords: Schistosomiasis, Chemoprophylaxis, Prevalence, Rural Populatio

Identification of a Second Mimicry Epitope from \u3ci\u3eAcanthamoeba castellanii\u3c/i\u3e that Induces CNS Autoimmunity by Generating Cross-Reactive T Cells for MBP 89–101 in SJL Mice

We had previously reported that Acanthamoeba castellanii (ACA) contains a mimicry epitope for proteolipid protein 139–151 capable of inducing central nervous system (CNS) autoimmunity in SJL/J mice. We now present evidence that ACA also contains a mimicry epitope for myelin basic protein (MBP) 89–101, a derivative from amoebic nicotinamide adenine dinucleotide dehydrogenase subunit 2 (NAD). The epitope, NAD 108–120, contains a discontinuous stretch of six amino acids in the core region (VVFFKNIILIGFL) sharing 46% identity with MBP 89–101 (VHFFKNIVTPRTP; identical residues are underlined). SJL mice immunized with NAD 108–120 develop encephalomyelitis similar to the disease induced by the cognate peptide. We demonstrate that NAD 108–120 induces T cells that cross-react with MBP 89–101; the antigen-sensitized T cells, which produce predominantly T helper (Th) 1 and Th17 cytokines, transfer disease in naive SJL recipients reminiscent of the disease induced with MBP 89–101. This is the first report to demonstrate that a solitary microbe can induce CNS autoimmunity by generating cross-reactive T cells for multiple myelin antigens