Histidine hydrogen bonding in MHC at pH 5 and pH 7 modeled by molecular docking and molecular dynamics simulations

Hydrogen bonds play important roles in maintaining the structure of proteins and in the formation of most biomolecular protein-ligand complexes. All amino acids can act as hydrogen bond donors and acceptors. Among amino acids, Histidine is unique, as it can exist in neutral or positively charged forms within the physiological pH range of 5.0 to 7.0. Histidine can thus interact with other aromatic residues as well as forming hydrogen bonds with polar and charged residues. The ability of His to exchange a proton lies at the heart of many important functional biomolecular interactions, including immunological ones. By using molecular docking and molecular dynamics simulation, we examine the influence of His protonation/deprotonation on peptide binding affinity to MHC class II proteins from locus HLA-DP. Peptide-MHC interaction underlies the adaptive cellular immune response, upon which the next generation of commercially-important vaccines will depend. Consistent with experiment, we find that peptides containing protonated His residues bind better to HLA-DP proteins than those with unprotonated His. Enhanced binding at pH 5.0 is due, in part, to additional hydrogen bonds formed between peptide His+ and DP proteins. In acidic endosomes, protein His79β is predominantly protonated. As a result, the peptide binding cleft narrows in the vicinity of His79β, which stabilizes the peptide - HLA-DP protein complex. © 2014 Bentham Science Publishers.

The Effect of Nutrient-Rich Effluents from Shrimp Farming on Mangrove Soil Carbon Storage and Geochemistry Under Semi-Arid Climate Conditions in Northern Brazil

A semi-arid mangrove estuary system in the northeast Brazilian coast (Ceará state) was selected for this study to (i) evaluate the impact of shrimp farm nutrient-rich wastewater effluents on the soil geochemistry and organic carbon (OC) storage and (ii) estimate the total amount of OC stored in mangrove soils (0–40 cm). Wastewater-affected mangrove forests were referred to as WAM and undisturbed areas as Non-WAM. Redox conditions and OC content were statistically correlated (P < 0.05) with seasonality and type of land use (WAM vs. Non-WAM). Eh values were from anoxic to oxic conditions in the wet season (from − 5 to 68 mV in WAM and from < 40 to > 400 mV in Non-WAM soils) and significantly higher (from 66 to 411 mV) in the dry season (P < 0.01). OC contents (0–40 cm soil depth) were significantly higher (P < 0.01) in the wet season than the dry season, and higher in Non-WAM soils than in WAM soils (values of 8.1 and 6.7 kg m− 2 in the wet and dry seasons, respectively, for Non-WAM, and values of 3.8 and 2.9 kg m− 2 in the wet and dry seasons, respectively, for WAM soils; P < 0.01). Iron partitioning was significantly dependent (P < 0.05) on type of land use, with a smaller degree of pyritization and lower Fe-pyrite presence in WAM soils compared to Non-WAM soils. Basal respiration of soil sediments was significantly influenced (P < 0.01) by type of land use with highest CO2 flux rates measured in the WAM soils (mean values of 0.20 mg CO2 h− 1–g− 1 C vs. 0.04 mg CO2 h− 1–g− 1 C). The OC storage reduction in WAM soils was potentially caused (i) by an increase in microbial activity induced by loading of nutrient-rich effluents and (ii) by an increase of strong electron acceptors [e.g., NO3−] that promote a decrease in pyrite concentration and hence a reduction in soil OC burial. The current estimated OC stored in mangrove soils (0–40 cm) in the state of Ceará is approximately 1 million t.

Methane and sulphate consumption in different habitats of the Håkon Mosby Mud Volcano (HMMV)

The Hakon Mosby Mud Volcano is a highly active methane seep hosting different chemosynthetic communities such as thiotrophic bacterial mats and siboglinid tubeworm assemblages. This study focuses on in situ measurements of methane fluxes to and from these different habitats, in comparison to benthic methane and oxygen consumption rates. By quantifying in situ oxygen, methane, and sulfide fluxes in different habitats, a spatial budget covering areas of 10-1000 -m diameter was established. The range of dissolved methane efflux (770-2 mmol m-2 d-1) from the center to the outer rim was associated with a decrease in temperature gradients from 46°C to < 1°C m-1, indicating that spatial variations in fluid flow control the distribution of benthic habitats and activities. Accordingly, total oxygen uptake (TOU) varied between the different habitats by one order of magnitude from 15 mmol m-2 d-1 to 161 mmol m-2 d-1. High fluid flow rates appeared to suppress benthic activities by limiting the availability of electron acceptors. Accordingly, the highest TOU was associated with the lowest fluid flow and methane efflux. This was most likely due to the aerobic oxidation of methane, which may be more relevant as a sink for methane as previously considered in submarine ecosystems.

Microbial Community Responses to Environmental Perturbation

Microorganisms mediate many biogeochemical processes critical to the functioning of ecosystems, which places them as an intermediate between environmental change and the resulting ecosystem response. Yet, we have an incomplete understanding of these relationships, how to predict them, and when they are influential. Understanding these dynamics will inform ecological principles developed for macroorganisms and aid expectations for microbial responses to new gradients. To address this research goal, I used two studies of environmental gradients and a literature synthesis.

With the gradient studies, I assessed microbial community composition in stream biofilms across a gradient of alkaline mine drainage. I used multivariate approaches to examine changes in the non-eukaryote microbial community composition of taxa (chapter 2) and functional genes (chapter 3). I found that stream biofilms at sites receiving alkaline mine drainage had distinct community composition and also differed in the composition of functional gene groups compared with unmined reference sites. Compositional shifts were not dominated by groups that could benefit from mining associated increases of terminal electron acceptors; two-thirds of responsive taxa and functional gene groups were negatively associated with mining. The majority of subsidies and stressors (nitrate, sulfate, conductivity) had no consistent relationships with taxa or gene abundances. However, methane metabolism genes were less abundant at mined sites and there was a strong, positive correlation between selenate reductase gene abundance and mining-associated selenium. These results highlighted the potential for indirect factors to also play an important role in explaining compositional shifts.

In the fourth chapter, I synthesized studies that use environmental perturbations to explore microbial community structure and microbial process connections. I examined nine journals (2009–13) and found that many qualifying papers (112 of 148) documented structure and process responses, but few (38 of 112 papers) reported statistically testing for a link. Of these tested links, 75% were significant. No particular approach for characterizing structure or processes was more likely to produce significant links. Process responses were detected earlier on average than responses in structure. Together, the findings suggested that few publications report statistically testing structure-process links; but when tested, links often occurred yet shared few commonalities in linked processes or structures and the techniques used for measuring them.

Although the research community has made progress, much work remains to ensure that the vast and growing wealth of microbial informatics data is translated into useful ecological information. In part, this challenge can be approached through using hypotheses to guide analyses, but also by being open to opportunities for hypothesis generation. The results from my dissertation work advise that it is important to carefully interpret shifts in community composition in relation to abiotic characteristics and recommend considering ecological, thermodynamic, and kinetic principles to understand the properties governing community responses to environmental perturbation.

Utilisation d’un modèle de co-culture cellulaire pour l’évaluation in vitro du transport inverse du cholestérol

De nouveaux modèles cellulaires in vitro par transfert de milieu et par coculture ont été mis au point afin d’évaluer la capacité des HDL à éliminer l’excès de cholestérol des tissus périphériques et de le transporter vers le foie afin d’être excrété par le foie, un processus nommé le transport inverse du cholestérol (TIC). Le système cellulaire par transfert in vitro où des macrophages J774 sont gorgés de LDL acétylées et marqués au 3H-cholestérol a été préalablement établi afin de mesurer par scintillation l’efflux de cholestérol marqué vers le milieu de culture contenant des accepteurs de cholestérol. Ce milieu conditionné est transféré sur des cellules HepG2 afin d’étudier l’influx du cholestérol marqué. Ce dernier nous permet d’observer un transport de cholestérol de 25 % hors des J774 et un transport de 39 000 cpm dans les HepG2 en utilisant un milieu contenant 2 % de sérums humains mis en commun. Une stimulation des cellules J774 par l’AMPc augmente l’efflux et l’influx d’environ 45 %. Des tests de preuve de concept ont été effectués sur le système cellulaire par co-culture qui utilise des chambres de Boyden où les J774 sont localisées au fond d’un puits et les HepG2 dans un insert, et où le milieu est partagé entre les deux types cellulaires. On a déterminé qu’une confluence densité de 60 000 cellules/cm2 sur un insert constitué d’une membrane de polyester avec des pores de 3,0 μm, sans autre revêtement, permet d’observer un influx spécifique au sérum d’environ 6 000 cpm associés aux cellules HepG2, où 50 % des comptes radioactifs sont dans les cellules et l’autre moitié présente à la surface cellulaire.

Geochemistry and geomicrobiology profiles of ferruginous sediment, Lake Towuti, Indonesia

Lake Towuti is a tectonic basin, surrounded by ultramafic rocks. Lateritic soils form through weathering and deliver abundant iron (oxy)hydroxides but very little sulfate to the lake and its sediment. To characterize the sediment biogeochemistry, we collected cores at three sites with increasing water depth and decreasing bottom water oxygen concentrations. Microbial cell densities were highest at the shallow site - a feature we attribute to the availability of labile organic matter and the higher abundance of electron acceptors due to oxic bottom water conditions. At the two other sites, OM degradation and reduction processes below the oxycline led to partial electron acceptor depletion. Genetic information preserved in the sediment as extracellular DNA provides information on aerobic and anaerobic heterotrophs related to Actinobacteria, Nitrospirae, Chloroflexi and Thermoplasmatales. These taxa apparently played a significant role in the degradation of sinking organic matter. However, extracellular DNA concentrations rapidly decrease with core depth. Despite very low sulfate concentrations, sulfate-reducing bacteria were present and viable in sediments at all three sites, as confirmed by measurement of potential sulfate reduction rates. Microbial community fingerprinting supported the presence of taxa related to Deltaproteobacteria and Firmicutes with demonstrated capacity for iron and sulfate reduction. Concomitantly, sequences of Ruminococcaceae, Clostridiales and Methanomicrobiales indicated potential for fermentative hydrogen and methane production. Such first insights into ferruginous sediments show that microbial populations perform successive metabolisms related to sulfur, iron and methane. In theory, iron reduction could reoxidize reduced sulfur compounds and desorb OM from iron minerals to allow remineralization to methane. Overall, we found that biogeochemical processes in the sediments can be linked to redox differences in the bottom waters of the three sites, like oxidant concentrations and the supply of labile OM. At the scale of the lacustrine record, our geomicrobiological study should provide a means to link the extant subsurface biosphere to past environments.

(Table 4) Distribution of bacterial populations in sediments of ODP Hole 128-798B

Sediment whole-round cores from a dedicated hole (798B) were obtained for detailed microbiological analysis, down to 518 m below the seafloor (mbsf). These sediments have characteristic bacterial profiles in the top 6 mbsf, with high but rapidly decreasing bacterial populations (total and dividing bacteria, and concentrations of different types of viable heterotrophic bacteria) and potential bacterial activities. Rates of thymidine incorporation into bacterial DNA and anaerobic sulfate reduction are high in the surface sediments and decrease rapidly down to 3 mbsf. Methanogenesis from CO2/H2 peaks below the maximum in sulfate reduction and although it decreases markedly down the core, is present at low rates at all but one depth. Consistent with these activities is the removal of pore-water sulfate, methane gas production, and accumulation of reduced sulfide species. Rates of decrease in bacterial populations slow down below 6 mbsf, and there are some distinct increases in bacterial populations and activities that continue over considerable depth intervals. These include a large and significant increase in total heterotrophic bacteria below 375 mbsf, which corresponds to an increase in the total bacterial population, bacterial viability, a small increase in potential rates of sulfate reduction, and the presence of thermogenic methane and other gases. Bacterial distributions seem to be controlled by the availability of terminal electron acceptors (e.g., sulfate), the bioavailability of organic carbon (which may be related to the dark/light bands within the sediment), and biological and geothermal methane production. Significant bacterial populations are present even in the deepest samples (518 mbsf) and hence it seems likely that bacteria may continue to be present and active much deeper than the sediments studied here. These results confirm and extend our previous results of bacterial activity within deep sediments of the Peru Margin from Leg 112, and to our knowledge this is the first comprehensive report of the presence of active bacterial populations from the sediment surface to in excess of 500 mbsf and sediments > 4 m.y. old.

Low temperature magneto-photoluminescence characterization of high purity gallium arsenide and indium phosphide

Low-temperature magneto-photoluminescence is a very powerful technique to characterize high purity GaAs and InP grown by various epitaxial techniques. These III-V compound semiconductor materials are used in a wide variety of electronic, optoelectronic and microwave devices. The large binding energy differences of acceptors in GaAs and InP make possible the identification of those impurities by low-temperature photoluminescence without the use of any magnetic field. However, the sensitivity and resolution provided by this technique rema1ns inadequate to resolve the minute binding energy differences of donors in GaAs and InP. To achieve higher sensitivity and resolution needed for the identification of donors, a magneto-photoluminescence system 1s installed along with a tunable dye laser, which provides resonant excitation. Donors 1n high purity GaAs are identified from the magnetic splittings of "two-electron" satellites of donor bound exciton transitions 1n a high magnetic field and at liquid helium temperature. This technique 1s successfully used to identify donors 1n n-type GaAs as well as 1n p-type GaAs in which donors cannot be identified by any other technique. The technique is also employed to identify donors in high purity InP. The amphoteric incorporation of Si and Ge impurities as donors and acceptors in (100), (311)A and (3ll)B GaAs grown by molecular beam epitaxy is studied spectroscopically. The hydrogen passivation of C acceptors in high purity GaAs grown by molecular beam epitaxy (MBE) and metalorganic chemical vapor deposition (MOCVD) 1s investigated using photoluminescence. Si acceptors ~n MBE GaAs are also found to be passivated by hydrogenation. The instabilities in the passivation of acceptor impurities are observed for the exposure of those samples to light. Very high purity MOCVD InP samples with extremely high mobility are characterized by both electrical and optical techniques. It is determined that C is not typically incorporated as a residual acceptor ~n high purity MOCVD InP. Finally, GaAs on Si, single quantum well, and multiple quantum well heterostructures, which are fabricated from III-V semiconductors, are also measured by low-temperature photoluminescence.

Crassaminicella profunda gen. nov., sp nov., an anaerobic marine bacterium isolated from deep-sea sediments

A novel, anaerobic, chemo-organotrophic bacterium, designated strain Ra1766HT, was isolated from sediments of the Guaymas basin (Gulf of California, Mexico) taken from a depth of 2002 m. Cells were thin, motile, Gram-stain-positive, flexible rods forming terminal endospores. Strain Ra1766H(T) grew at temperatures of 25-45 degrees C (optimum 30 degrees C), pH 6.7-8.1 (optimum 7.5) and in a salinity of 5-60 g l(-1) NaCl (optimum 30 g l(-1)). It was an obligate heterotrophic bacterium fermenting carbohydrates (glucose and mannose) and organic acids (pyruvate and succinate). Casamino acids and amino acids (glutamate, aspartate and glycine) were also fermented. The main end products from glucose fermentation were acetate, butyrate, ethanol, H-2 and CO2. Sulfate, sulfite, thiosulfate, elemental sulfur, fumarate, nitrate, nitrite and Fe(III) were not used as terminal electron acceptors. The predominant cellular fatty acids were C-14 : 0, C-16:1 omega 7, C-16:1 omega 7 DMA and C-16:0. The main polar lipids consisted of phosphatidylglycerol, diphosphatidylglycerol, phosphatidylethanolamine and phospholipids. The G +C content of the genomic DNA was 33.7 molo/o. Phylogenetic analysis of the 16S rRNA gene sequence indicated that strain Ra1766H(T) was affiliated to cluster XI of the order Clostridia les, phylum Firmicutes. The closest phylogenetic relative of Ra1766H(T) was Geosporobacter subterraneus (94.2% 16S rRNA gene sequence similarity). On the basis of phylogenetic inference and phenotypic properties, strain Ra1766H(T) (=DSM 27501(T)=JCM 19377(T)) is proposed to be the type strain of a novel species of a novel genus, named Crassaminicella pro funda.

Utilisation d’un modèle de co-culture cellulaire pour l’évaluation in vitro du transport inverse du cholestérol

De nouveaux modèles cellulaires in vitro par transfert de milieu et par coculture ont été mis au point afin d’évaluer la capacité des HDL à éliminer l’excès de cholestérol des tissus périphériques et de le transporter vers le foie afin d’être excrété par le foie, un processus nommé le transport inverse du cholestérol (TIC). Le système cellulaire par transfert in vitro où des macrophages J774 sont gorgés de LDL acétylées et marqués au 3H-cholestérol a été préalablement établi afin de mesurer par scintillation l’efflux de cholestérol marqué vers le milieu de culture contenant des accepteurs de cholestérol. Ce milieu conditionné est transféré sur des cellules HepG2 afin d’étudier l’influx du cholestérol marqué. Ce dernier nous permet d’observer un transport de cholestérol de 25 % hors des J774 et un transport de 39 000 cpm dans les HepG2 en utilisant un milieu contenant 2 % de sérums humains mis en commun. Une stimulation des cellules J774 par l’AMPc augmente l’efflux et l’influx d’environ 45 %. Des tests de preuve de concept ont été effectués sur le système cellulaire par co-culture qui utilise des chambres de Boyden où les J774 sont localisées au fond d’un puits et les HepG2 dans un insert, et où le milieu est partagé entre les deux types cellulaires. On a déterminé qu’une confluence densité de 60 000 cellules/cm2 sur un insert constitué d’une membrane de polyester avec des pores de 3,0 μm, sans autre revêtement, permet d’observer un influx spécifique au sérum d’environ 6 000 cpm associés aux cellules HepG2, où 50 % des comptes radioactifs sont dans les cellules et l’autre moitié présente à la surface cellulaire.

Identification and characterization of shallow impurity states in gallium arsenide and indium phosphide using photothermal ionization spectroscopy

A model of far infrared (FIR) dielectric response of shallow impurity states in a semiconductor has been developed and is presented for the specific case of the shallow donor transitions in high purity epitaxial GaAs. The model is quite general, however, and should be applicable with slight modification, not only to shallow donors in other materials such as InP, but also to shallow acceptors and excitons. The effects of the enormous dielectric response of shallow donors on the FIR optical properties of reflectance, transmittance, and absorptance, and photoconductive response of high purity epitaxial GaAs films are predicted and compared with experimental photothermal ionization spectra. The model accounts for many of the peculiar features that are frequently observed in these spectra, one of which was the cause of erroneous donor identifications in the early doping experiments. The model also corrects some commonly held misconceptions concerning photo-thermal ionization peak widths and amplitudes and their relationships to donor and acceptor concentrations. These corrections are of particular relevance to the proper interpretation of photothermal ionization spectra in the study of impurity incorporation in high purity epitaxial material. The model also suggests that the technique of FIR reflectance, although it has not been widely employed, should be useful in the study of shallow impurities in semiconductors.

Maan korvessa kulkevi – Evakkolasten kasvukokemusten tunnemuistot ja hyvinvointi

Tutkimus tarkastelee luovutetusta Karjalasta lähtöisin olevien evakkolasten tunnemuistoja ja niiden yhteyttä myöhempään hyvinvointiin. Keskiössä ovat koti- ja koulukasvatusta muistelevien ikäihmisten kokemukset. Tutkimus on kasvatushistoriallinen ja tieteidenvälinen. Se liikkuu historian sekä yhteiskunta- ja käyttäytymistieteiden rajapinnoilla. Aineiston 631 kyselyvastausta ja 11 haastattelua on analysoitu monimetodisesti tilastollisen ja sisällön analyysin avulla. Tulosten mukaan ikääntyvät evakkolapset voivat pääsääntöisesti hyvin. He ovat kotikasvatuksessaan omaksuneet vahvat selviytymis- ja sopeutumistaidot sekä sisukkuuden. Kodin kasvatusilmapiiri on menetyksistä huolimatta koettu turvalliseksi. Kasvatuksessa omaksutuissa sananlaskuissa korostuu vahvimmin tulevaisuuden toivosta kiinni pitäminen. Kasvatusmetodit myötäilevät suomalaista kasvatustraditiota, jossa lapsia on enimmäkseen palkittu hyvästä työstä sanallisesti ja rangaistu tottelemattomuudesta ruumiillisesti. Surua ja menetyksiä on käsitelty vaikenemalla. Yhdeksi elämää kannattelevaksi voimaksi ovat osoittautuneet laulut, joista suosituin oli Maan korvessa kulkevi lapsosen tie. Kasvatusperintönä evakkolapset haluavat jättää mallin työteliäästä, rehellisestä, toiset huomioivasta ja juuristaan tietoisesta kansalaisesta. Koulukokemusten osalta tutkimus osoittaa kantaväestöstä poikkeavan uskonnollisen ja kielellisen taustan olleen merkittävin syrjityksi tulemisen syy. Ortodoksitaustaisilla lapsilla oli muita evakkolapsia enemmän kiusaamiskokemuksia. Metaforien avulla kuvatuista tunnemuistoista löytyi kuusi erilaista selviytyjätyyppiä: selviytyjäsankarit, identiteetin etsijät, optimistiset toimijat, trauman työstäjät, kohtalonsa hyväksyjät ja menetysten kantajat. Valtaosa evakkolapsista kuuluu positiivista tunnemuistoa kantavien ryhmään. Kielteisillä tunnemuistoilla on tilastollisesti merkitsevä negatiivinen yhteys ikääntyvän hyvinvointiin. Tutkimus lisää ymmärrystä evakkolasten kasvukokemuksista ja tarjoaa uutta tietoa heidän selviytymisestään ja hyvinvoinnistaan. Tutkimus osoittaa evakkolasten tasapainoisen kotikasvatuksen, vastavuoroisten vertaissuhteiden ja suositun oppilaan aseman vahvistavan yhteisöön kuulumista ja sosiaalista pystyvyyttä sekä olevan hyvinvoinnin positiivisia tekijöitä. Kodin melankolisuudella, eristyneisyydellä ja koulukiusaamisella on negatiivinen yhteys hyvinvointiin. Tuloksia voidaan soveltaa nykypäivän pakolaislasten kotouttamiseen, jossa ensiarvoisen tärkeää on perheiden osallisuuden ja integroitumisen tukeminen uudessa ympäristössä. Tämän onnistumiseksi on tärkeää huomioida lasten yksilölliset selviytymisstrategiat ja käsitellä kodin menetystraumaa.

Development of two charge transfer complex spectrophotometric methods for determination of tofisopam in tablet dosage form

Purpose: To develop a simple, fast and sensitive spectrophotometric method for the determination of tofisopam in tablet dosage form. Methods: Tofisopam as n-electron donor was reacted with two π-acceptors, namely, chloranilic acid (ChA), and 7,7,8,8 tetracyanoquinodimethane (TCNQ) to form charge transfer complexes. The complexes were evaluated spectrophotometrically at 520 and 824 nm for ChA and TCNQ, respectively. The optimum conditions for the reaction were determined and optimized. The developed method was compared with Japanese Pharmacopeia method. Results: The calibration curve was linear in the ranges 25 – 125 and 30 – 150 μg/mL for ChA and TCNQ, respectively. The lower limit of detection was 8.0 and 10.0 μg/mL for ChA and TCNQ, respectively while the slope and intercept of the calibration curves were 0.0025 and 0.011 and 0.0115 and -0.237, for ChA and TCNQ, respectively. Conclusion: The developed methods for tofisopam have good accuracy and precision, and comparable to a standard pharmacopeial method. The methods can be applied for routine analysis and in quality control.

DEVELOPMENT OF A 2-DIMENSIONAL FINITE VOLUME MODEL TO ASSESS HYDRODYNAMIC AND MICROBIAL CONTROLS ON DNAPL DISSOLUTION AND DETOXIFICATION

Tetrachloroethene (PCE) and trichloroethene (TCE) form dense non-aqueous phase liquids (DNAPLs), which are persistent groundwater contaminants. DNAPL dissolution can be "bioenhanced" via dissolved contaminant biodegradation at the DNAPL-water interface. This research hypothesized that: (1) competitive interactions between different dehalorespiring strains can significantly impact the bioenhancement effect, and extent of PCE dechlorination; and (2) hydrodynamics will affect the outcome of competition and the potential for bioenhancement and detoxification. A two-dimensional coupled flowtransport model was developed, with a DNAPL pool source and multiple microbial species. In the scenario presented, Dehalococcoides mccartyi 195 competes with Desulfuromonas michiganensis for the electron acceptors PCE and TCE. Simulations under biostimulation and low velocity (vx) conditions suggest that the bioenhancement with Dsm. michiganensis alone was modestly increased by Dhc. mccartyi 195. However, the presence of Dhc. mccartyi 195 enhanced the extent of PCE transformation. Hydrodynamic conditions impacted the results by changing the dominant population under low and high vx conditions.