The Structure and Functions of Hantavirus Nucleocapsid Protein

Hantaviruses, members of the genus Hantavirus in the Bunyaviridae family, are enveloped single-stranded RNA viruses with tri-segmented genome of negative polarity. In humans, hantaviruses cause two diseases, hemorrhagic fever with renal syndrome (HFRS) and hantavirus pulmonary syndrome (HPS), which vary in severity depending on the causative agent. Each hantavirus is carried by a specific rodent host and is transmitted to humans through excreta of infected rodents. The genome of hantaviruses encodes four structural proteins: the nucleocapsid protein (N), the glycoproteins (Gn and Gc), and the polymerase (L) and also the nonstructural protein (NSs). This thesis deals with the functional characterization of hantavirus N protein with regard to its structure. Structural studies of the N protein have progressed slowly and the crystal structure of the whole protein is still not available, therefore biochemical assays coupled with bioinformatical modeling proved essential for studying N protein structure and functions. Presumably, during RNA encapsidation, the N protein first forms intermediate trimers and then oligomers. First, we investigated the role of N-terminal domain in the N protein oligomerization. The results suggested that the N-terminal region of the N protein forms a coiled-coil, in which two antiparallel alpha helices interact via their hydrophobic seams. Hydrophobic residues L4, I11, L18, L25 and V32 in the first helix and L44, V51, L58 and L65 in the second helix were crucial for stabilizing the structure. The results were consistent with the head-to-head, tail-to-tail model for hantavirus N protein trimerization. We demonstrated that an intact coiled-coil structure of the N terminus is crucial for the oligomerization capacity of the N protein. We also added new details to the head-to-head, tail-to-tail model of trimerization by suggesting that the initial step is based on interaction(s) between intact intra-molecular coiled-coils of the monomers. We further analyzed the importance of charged aa residues located within the coiled-coil for the N protein oligomerization. To predict the interacting surfaces of the monomers we used an upgraded in silico model of the coiled-coil domain that was docked into a trimer. Next the predicted target residues were mutated. The results obtained using the mammalian two-hybrid assay suggested that conserved charged aa residues within the coiled-coil make a substantial contribution to the N protein oligomerization. This contribution probably involves the formation of interacting surfaces of the N monomers and also stabilization of the coiled-coil via intramolecular ionic bridging. We proposed that the tips of the coiled-coils are the first to come into direct contact and thus initiate tight packing of the three monomers into a compact structure. This was in agreement with the previous results showing that an increase in ionic strength abolished the interaction between N protein molecules. We also showed that residues having the strongest effect on the N protein oligomerization are not scattered randomly throughout the coiled-coil 3D model structure, but form clusters. Next we found evidence for the hantaviral N protein interaction with the cytoplasmic tail of the glycoprotein Gn. In order to study this interaction we used the GST pull-down assay in combination with mutagenesis technique. The results demonstrated that intact, properly folded zinc fingers of the Gn protein cytoplasmic tail as well as the middle domain of the N protein (that includes aa residues 80 248 and supposedly carries the RNA-binding domain) are essential for the interaction. Since hantaviruses do not have a matrix protein that mediates the packaging of the viral RNA in other negatve stranded viruses (NSRV), hantaviral RNPs should be involved in a direct interaction with the intraviral domains of the envelope-embedded glycoproteins. By showing the N-Gn interaction we provided the evidence for one of the crucial steps in the virus replication at which RNPs are directed to the site of the virus assembly. Finally we started analysis of the N protein RNA-binding region, which is supposedly located in the middle domain of the N protein molecule. We developed a model for the initial step of RNA-binding by the hantaviral N protein. We hypothesized that the hantaviral N protein possesses two secondary structure elements that initiate the RNA encapsidation. The results suggest that amino acid residues (172-176) presumably act as a hook to catch vRNA and that the positively charged interaction surface (aa residues 144-160) enhances the initial N-RNA interacation. In conclusion, we elucidated new functions of hantavirus N protein. Using in silico modeling we predicted the domain structure of the protein and using experimental techniques showed that each domain is responsible for executing certain function(s). We showed that intact N terminal coiled-coil domain is crucial for oligomerization and charged residues located on its surface form a interaction surface for the N monomers. The middle domain is essential for interaction with the cytoplasmic tail of the Gn protein and RNA binding.

Temperature dependence of g values in tetragonal ionic Cu++ salts

A temperature dependence has been observed in the spin-Hamiltonian parameters of the Cu++ ion in a tetragonal crystal field and the variation has been interpreted in terms of vibronic effects.

First simultaneous measurement of the top quark mass in the lepton+jets and dilepton channels at CDF

We present a measurement of the mass of the top quark using data corresponding to an integrated luminosity of 1.9fb^-1 of ppbar collisions collected at sqrt{s}=1.96 TeV with the CDF II detector at Fermilab's Tevatron. This is the first measurement of the top quark mass using top-antitop pair candidate events in the lepton + jets and dilepton decay channels simultaneously. We reconstruct two observables in each channel and use a non-parametric kernel density estimation technique to derive two-dimensional probability density functions from simulated signal and background samples. The observables are the top quark mass and the invariant mass of two jets from the W decay in the lepton + jets channel, and the top quark mass and the scalar sum of transverse energy of the event in the dilepton channel. We perform a simultaneous fit for the top quark mass and the jet energy scale, which is constrained in situ by the hadronic W boson mass. Using 332 lepton + jets candidate events and 144 dilepton candidate events, we measure the top quark mass to be mtop=171.9 +/- 1.7 (stat. + JES) +/- 1.1 (syst.) GeV/c^2 = 171.9 +/- 2.0 GeV/c^2.

First Observation of Vector Boson Pairs in a Hadronic Final State at the Tevatron Collider

We present the first observation in hadronic collisions of the electroweak production of vector boson pairs (VV, V=W, Z) where one boson decays to a dijet final state. The data correspond to 3.5  fb-1 of integrated luminosity of pp̅ collisions at √s=1.96  TeV collected by the CDF II detector at the Fermilab Tevatron. We observe 1516±239(stat)±144(syst) diboson candidate events and measure a cross section σ(pp̅ →VV+X) of 18.0±2.8(stat)±2.4(syst)±1.1(lumi)  pb, in agreement with the expectations of the standard model.

First Observation of Vector Boson Pairs in a Hadronic Final State at the Tevatron Collider

We present the first observation in hadronic collisions of the electroweak production of vector boson pairs (VV, V=W,Z) where one boson decays to a dijet final state . The data correspond to 3.5 inverse femtobarns of integrated luminosity of ppbar collisions at sqrt(s)=1.96 TeV collected by the CDFII detector at the Fermilab Tevatron. We observe 1516+/-239(stat)+/-144(syst) diboson candidate events and measure a cross section sigma(ppbar->VV+X) of 18.0+/-2.8(stat)+/-2.4(syst)+/-1.1(lumi) pb, in agreement with the expectations of the standard model.

Raman spectra of crystalline double sulphates

Raman spectra of single crystals of K2M(SO4)2 · 6 H2O where M=Mg, Zn, Ni or Co have been recorded for the first time using λ 2537 as the exciting radiation. The corresponding five single sulphates have also been studied. Interesting results concerning the substitution of magnesium, zinc, nickel or cobalt in the double sulphate lattice on the sulphate frequencies are observed. The lattice spectra of these double sulphates are analysed group theoretically and discussed in relation to the lattice spectra of the corresponding individual sulphates. Certain new results concerning the Raman spectra of the individual sulphates have also been obtained and in the case of CoSO4 · 7 H2O the spectrum has been recorded for the first time.

Raman spectra of single crystals of dicarboxylic acids - Part I. β-malonic acid and β-succinic acid

Raman spectra of single crystals of β-malonic acid and β-succinic acid have been photographed using λ 2536·5 radiation. 32 Raman lines have been recorded in the case of β-malonic acid. Of these 21 lines have been recorded for the first time. The three intense lattice lines at 52, 90 and 144 cm.-1 have been attributed to rotational lattice oscillations. 29 Raman lines in the case of β-succinic acid have been recorded. The entire lattice spectrum and many internal frequencies have been recorded for the first time. The three intense lattice lines at 80, 135 and 160 cm.-1 have been assigned to the rotational oscillations of the two molecules of the succinic acid in the unit cell.

Molecular Basis for the Role of Staphylococcus aureus Penicillin Binding Protein 4 in Antimicrobial Resistance

Penicillin binding proteins (PBPs) are membrane-associated proteins that catalyze the final step of murein biosynthesis. These proteins function as either transpeptidases or carboxypeptidases and in a few cases demonstrate transglycosylase activity. Both transpeptidase and carboxypeptidase activities of PBPs occur at the D-Ala-D-Ala terminus of a murein precursor containing a disaccharide pentapeptide comprising N-acetyl-glucosamine and N-acetyl-muramic acid-L-Ala-D-Glu-L-Lys-D-Ala-D-Ala. beta-Lactam antibiotics inhibit these enzymes by competing with the pentapeptide precursor for binding to the active site of the enzyme. Here we describe the crystal structure, biochemical characteristics, and expression profile of PBP4, a low-molecular-mass PBP from Staphylococcus aureus strain COL. The crystal structures of PBP4-antibiotic complexes reported here were determined by molecular replacement, using the atomic coordinates deposited by the New York Structural Genomics Consortium. While the pbp4 gene is not essential for the viability of S. aureus, the knockout phenotype of this gene is characterized by a marked reduction in cross-linked muropeptide and increased vancomycin resistance. Unlike other PBPs, we note that expression of PBP4 was not substantially altered under different experimental conditions, nor did it change across representative hospital- or community-associated strains of S. aureus that were examined. In vitro data on purified recombinant S. aureus PBP4 suggest that it is a beta-lactamase and is not trapped as an acyl intermediate with beta-lactam antibiotics. Put together, the expression analysis and biochemical features of PBP4 provide a framework for understanding the function of this protein in S. aureus and its role in antimicrobial resistance.

Hur höra tyst kunskap? Utveckling av en metod för studier av tyst kunnande

Det tysta kunnandet utgör en stor del av kunskapsresursen både hos oss som individer och i arbetsorganisationerna. Trots att vi omger oss med böcker, manualer och databaser, som alla är exempel på explicit kunskap, så är det ”den rätta känslan”, erfarenheten och våra färdigheter som avgör om och hur vi klarar av våra uppgifter. Dessa begrepp är alla relaterade till den tysta dimensionen av kunskap. En dimension som traditionellt karaktäriserats som abstrakt, individuell, omedveten, praktisk, erfarenhetsbaserad och framför allt svår att uttrycka. Alla dessa är karaktärsdrag som ställt speciella krav inom kunskapsforskning och -ledning. Resultatet av detta är att både forskning och ledning av det tysta kunnandet har åsidosatts till förmån för forskning och ledning av explicit kunskap. Ett bidragande problem har varit bristen på lämpliga metoder för att ur ett företagsekonomiskt perspektiv studera och leda tyst kunnande. Ett annat problem har varit oklarhet i begreppet tyst kunskap. Detta har lett till brist på förståelse och/eller missförstånd. För att råda bot på svårigheten att uttrycka vårt tysta kunnande har människan utvecklat olika begrepp som i vår vardagskommunikation symboliserar tyst kunnande. Begrepp som intuition, människokännedom, förhandlingsförmåga och kultur används vanligt och med dem uttrycker vi den tysta dimensionen av kunnande. Dessa begrepp utgör även grunden för den intervjumetod som utvecklats för att empiriskt studera eller i ledningssyfte kartlägga tyst kunnande. Metoden använder dessa ”Epitet för Tyst Kunskap” (ETK) som bas för stimuluskort-intervjuer. Intervjuer som visat sig öka möjligheten att utforska och kartlägga tyst kunnande i organisationer oberoende av om man är forskare eller företagsledare.

ICT and the Shaping of Society: Exploring Human -ICT Relationships in Everyday Life (summary section only)

This thesis explores the relationship between humans and ICTs (information and communication technologies). As ICTs are increasingly penetrating all spheres of social life, their role as mediators – between people, between people and information, and even between people and the natural world – is expanding, and they are increasingly shaping social life. Yet, we still know little of how our life is affected by their growing role. Our understanding of the actors and forces driving the accelerating adoption of new ICTs in all areas of life is also fairly limited. This thesis addresses these problems by interpretively exploring the link between ICTs and the shaping of society at home, in the office, and in the community. The thesis builds on empirical material gathered in three research projects, presented in four separate essays. The first project explores computerized office work through a case study. The second is a regional development project aiming at increasing ICT knowledge and use in 50 small-town families. In the third, the second project is compared to three other longitudinal development projects funded by the European Union. Using theories that consider the human-ICT relationship as intertwined, the thesis provides a multifaceted description of life with ICTs in contemporary information society. By oscillating between empirical and theoretical investigations and balancing between determinist and constructivist conceptualisations of the human-ICT relationship, I construct a dialectical theoretical framework that can be used for studying socio-technical contexts in society. This framework helps us see how societal change stems from the complex social processes that surround routine everyday actions. For example, interacting with and through ICTs may change individuals’ perceptions of time and space, social roles, and the proper ways to communicate – changes which at some point in time result in societal change in terms of, for example, new ways of acting and knowing things.

Maximum Loss Calculation using Scenario Analysis, Heavy Tails and Implied Volatility Patterns

The objective of this paper is to improve option risk monitoring by examining the information content of implied volatility and by introducing the calculation of a single-sum expected risk exposure similar to the Value-at-Risk. The figure is calculated in two steps. First, there is a need to estimate the value of a portfolio of options for a number of different market scenarios, while the second step is to summarize the information content of the estimated scenarios into a single-sum risk measure. This involves the use of probability theory and return distributions, which confronts the user with the problems of non-normality in the return distribution of the underlying asset. Here the hyperbolic distribution is used to describe one alternative for dealing with heavy tails. Results indicate that the information content of implied volatility is useful when predicting future large returns in the underlying asset. Further, the hyperbolic distribution provides a good fit to historical returns enabling a more accurate definition of statistical intervals and extreme events.

Hormones andCuscuta development: In vitro induction of haustoria by cytokinin and its inhibition by other hormones

Cytokinins induced haustoria formation in excised 10-mm segments ofCuscuta vine, the subapical 25-to-50-mm region being most responsive, producing a mean of 4–6 haustoria per segment. The order of effectiveness of cytokinins continuously applied (72 h) was 6-benzylaminopurine (BA) ges isopentenyladenine (iP) Gt zeatin (Z). Ribosides of BA and Z were as effective as the bases, whereas riboside of iP ([9R]iP) was half as effective as iP. Haustoria induction was influenced by weather and seasonal conditions at the time of vine collection; materials obtained on warm, sunny days responded better than those obtained on rainy, cloudy, or cool days. Haustoria were induced equally well all around the segment, and no thigmostimulus was needed for induction. p ]A 10-min pulse of 100 mgrM BA induced half as many haustoria as a 60-min pulse or continuous application of BA. White light inhibited haustoria induction elicited by a short (30-min) pulse of BA, whereas a longer (120-min) BA application overcame this light inhibition. Auxins (IAA or NAA, 1–10 mgrM), gibberellin (GA3, 1–10 mgrM), ethylene (as ethrel, 10–100 mgrM), and abscisic acid (ABA, 100 mgrM) were individually inhibitory (60–80%) with respect to haustoria induction when given continuously with 50 mgrM BA. A 60-min pulse of auxins (10 mgrM), GA3 (100 mgrM), or ethrel (10 mgrM), given at various time intervals during or after a 60-min pulse of 100 mgrM BA, showed that inhibition was maximal (70–95%) between 4 and 16 h of BA application and negligible (GA3) or much reduced (auxin, ethrel) at 20 h, indicating a ldquocommitmentrdquo to haustoria formation by this time.

Nota sullo status della gallina prataiola (Tetrax tetrax) nella Zona di Protezione Speciale Piana di Semestene, Bonorva, Macomer e Bortigali (Sardegna), 2007-2009

The little bustard Tetrax tetrax is a bird of agro-pastoral landscapes rapidly disappearing from most of Western Europe due to agricultural intensification. In Italy, the species is virtually extinct from the mainland but still occurs in Sardinia, where four Special Protection Areas (SPAs) have been designated in 2007 to protect the bustard and related habitat. In this note, we document a steep decline (between 50 and 87,5%) of the species during the last decade in one of those four SPAs, Plains of Semestene, Bonorva, Macomer and Bortigali. However, during summer 2009, a group of 17 individuals was found within the SPA, confirming that the species, although at low numbers (5-15 displaying males), still breeds successfully in the area. Potential limiting factors are also discussed. We urge to undertake conservation measures based on solid scientific evidence if the local population is to be saved from probable extinction in the near future.