Update of the Gene Discovery Program in Schistosoma mansoni with the Expressed Sequence Tag Approach

Continuing the Schistosoma mansoni Genome Project 363 new templates were sequenced generating 205 more ESTs corresponding to 91 genes. Seventy four of these genes (81%) had not previously been described in S. mansoni. Among the newly discovered genes there are several of significant biological interest such as synaptophysin, NIFs-like and rho-GDP dissociation inhibitor

Purification of recombinant proteins by chemical removal of the affinity tag.

The efficient removal of a N- or C-terminal purification tag from a fusion protein is necessary to obtain a protein in a pure and active form, ready for use in human or animal medicine. Current techniques based on enzymatic cleavage are expensive and result in the presence of additional amino acids at either end of the proteins, as well as contaminating proteases in the preparation. Here we evaluate an alternative method to the one-step affinity/protease purification process for large-scale purification. It is based upon the cyanogen bromide (CNBr) cleavage at a single methionine placed in between a histidine tag and a Plasmodium falciparum antigen. The C-terminal segment of the circumsporozoite polypeptide was expressed as a fusion protein with a histidine tag in Escherichia coli purified by Ni-NAT agarose column chromatography and subsequently cleaved by CNBr to obtain a polypeptide without any extraneous amino acids derived from the cleavage site or from the affinity purification tag. Thus, a recombinant protein is produced without the need for further purification, demonstrating that CNBr cleavage is a precise, efficient, and low-cost alternative to enzymatic digestion, and can be applied to large-scale preparations of recombinant proteins.

Monitoring of NY-ESO-1 specific CD4+ T cells using molecularly defined MHC class II/His-tag-peptide tetramers.

MHC-peptide tetramers have become essential tools for T-cell analysis, but few MHC class II tetramers incorporating peptides from human tumor and self-antigens have been developed. Among limiting factors are the high polymorphism of class II molecules and the low binding capacity of the peptides. Here, we report the generation of molecularly defined tetramers using His-tagged peptides and isolation of folded MHC/peptide monomers by affinity purification. Using this strategy we generated tetramers of DR52b (DRB3*0202), an allele expressed by approximately half of Caucasians, incorporating an epitope from the tumor antigen NY-ESO-1. Molecularly defined tetramers avidly and stably bound to specific CD4(+) T cells with negligible background on nonspecific cells. Using molecularly defined DR52b/NY-ESO-1 tetramers, we could demonstrate that in DR52b(+) cancer patients immunized with a recombinant NY-ESO-1 vaccine, vaccine-induced tetramer-positive cells represent ex vivo in average 1:5,000 circulating CD4(+) T cells, include central and transitional memory polyfunctional populations, and do not include CD4(+)CD25(+)CD127(-) regulatory T cells. This approach may significantly accelerate the development of reliable MHC class II tetramers to monitor immune responses to tumor and self-antigens.

Effects of weight loss and exercise on insulin resistance, and intramyocellular triacylglycerol, diacylglycerol and ceramide.

AIMS/HYPOTHESIS: Intramyocellular lipids, including diacylglycerol (DAG) and ceramides, have been linked to insulin resistance. This randomised repeated-measures study examined the effects of diet-induced weight loss (DIWL) and aerobic exercise (EX) on insulin sensitivity and intramyocellular triacylglycerol (IMTG), DAG and ceramide. METHODS: Sixteen overweight to obese adults (BMI 30.6 ± 0.8; 67.2 ± 4.0 years of age) with either impaired fasting glucose, or impaired glucose tolerance completed one of two lifestyle interventions: DIWL (n = 8) or EX (n = 8). Insulin sensitivity was determined using hyperinsulinaemic-euglycaemic clamps. Intramyocellular lipids were measured in muscle biopsies using histochemistry and tandem mass spectrometry. RESULTS: Insulin sensitivity was improved with DIWL (20.6 ± 4.7%) and EX (19.2 ± 12.9%). Body weight and body fat were decreased by both interventions, with greater decreases in DIWL compared with EX. Muscle glycogen, IMTG content and oxidative capacity were all significantly (p < 0.05) decreased with DIWL and increased with EX. There were decreases in DAG with DIWL (-12.4 ± 14.6%) and EX (-40.9 ± 12.0%). Ceramide decreased with EX (-33.7 ± 11.2%), but not with DIWL. Dihydroceramide was decreased with both interventions. Sphingosine was decreased only with EX. Changes in total DAG, total ceramides and other sphingolipids did not correlate with changes in glucose disposal. Stearoyl-coenzyme A desaturase 1 (SCD1) content was decreased with DIWL (-19.5 ± 8.5%, p < 0.05), but increased with EX (19.6 ± 7.4%, p < 0.05). Diacylglycerol acyltransferase 1 (DGAT1) was unchanged with the interventions. CONCLUSIONS/INTERPRETATION: Diet-induced weight loss and exercise training both improved insulin resistance and decreased DAG, while only exercise decreased ceramides, despite the interventions having different effects on IMTG. These alterations may be mediated through differential changes in skeletal muscle capacity for oxidation and triacylglycerol synthesis. TRIAL REGISTRATION: ClinicalTrials.gov NCT00766298.

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Poster at Open Repositories 2014, Helsinki, Finland, June 9-13, 2014

Reversible flow of cholesteryl ester between high-density lipoproteins and triacylglycerol-rich particles is modulated by the fatty acid composition and concentration of triacylglycerols

We determined the influence of fasting (FAST) and feeding (FED) on cholesteryl ester (CE) flow between high-density lipoproteins (HDL) and plasma apoB-lipoprotein and triacylglycerol (TG)-rich emulsions (EM) prepared with TG-fatty acids (FAs). TG-FAs of varying chain lengths and degrees of unsaturation were tested in the presence of a plasma fraction at d > 1.21 g/mL as the source of CE transfer protein. The transfer of CE from HDL to FED was greater than to FAST TG-rich acceptor lipoproteins, 18% and 14%, respectively. However, percent CE transfer from HDL to apoB-containing lipoproteins was similar for FED and FAST HDL. The CE transfer from HDL to EM depended on the EM TG-FA chain length. Furthermore, the chain length of the monounsaturated TG-containing EM showed a significant positive correlation of the CE transfer from HDL to EM (r = 0.81, P < 0.0001) and a negative correlation from EM to HDL (r = -041, P = 0.0088). Regarding the degree of EM TG-FAs unsaturation, among EMs containing C18, the CE transfer was lower from HDL to C18:2 compared to C18:1 and C18:3, 17.7%, 20.7%, and 20%, respectively. However, the CE transfer from EMs to HDL was higher to C18:2 than to C18:1 and C18:3, 83.7%, 51.2%, and 46.3%, respectively. Thus, the EM FA composition was found to be the rate-limiting factor regulating the transfer of CE from HDL. Consequently, the net transfer of CE between HDL and TG-rich particles depends on the specific arrangement of the TG acyl chains in the lipoprotein particle core.

Elämä omin kuvin ja sanoin. Omaelämäkerrallisuus, identiteetti ja subjekti Ulli Lustin sarjakuvassa Heute ist der letzte Tag vom Rest deines Lebens

Ulli Lustin omaelämäkerrallinen Heute ist der letzte Tag vom Rest deines Lebens (2009) (suom. Tänään on loppuelämäsi viimeinen päivä, 2013) on 2000-luvun kansainvälisesti menestynein saksankielinen sarjakuva. 1980-luvulle sijoittuva teos kuvaa nuoren punkkari-Ullin vaiheikasta matkaa Wienistä Italiaan ja sen halki. Matkakertomus syvenee kasvukertomukseksi, sillä matkan tapahtumat saavat Ullin tarkastelemaan maailmaa ja itseään uusin silmin. Teos edustaa taidesarjakuvaa viime vuosikymmeninä hallinnutta elämäkerrallisen ja dokumentaarisen sarjakuvan kautta. Fiktion rajojen ylittäminen on nähty pyrkimyksenä laajentaa sarjakuvan ilmaisurepertuaaria ja vahvistaa sarjakuvan arvostusta kulttuurin kentällä. Ei-fiktiivisen sarjakuvan aalto on vaikuttanut myös sarjakuvatutkimukseen, jonka piirissä elämäkerrallisuus ja dokumentaarisuus ovat tämän hetken tutkituimpia ilmiöitä. Tutkielmani asettuu osaksi omaelämäkerrallisen sarjakuvan tutkimusta, mutta pyrin huomioimaan analyysissani myös modernin omaelämäkerran laajemman, aina modernin ajan alkuun ulottuvan kirjoittamisen ja tutkimisen perinteen. Tarkastelen erityisesti sitä, miten teos kertoo omaelämäkerrallisen tarinan ja miten se käsittelee omaelämäkertalajille keskeisiä identiteetin, itseymmärryksen ja autonomisen toimijuuden kysymyksiä. Yhdistän sarjakuvailmaisun analyysin tutkielmassani kulttuurihistorialliseen ja sosiologiseen otteeseen. Ymmärrän sarjakuvan omalakisena ilmaisumuotonaan, mutta hyödynnän analyysissani tarpeen mukaan myös kirjallisuuden ja elokuvan tutkimusta. Osoitan tutkielmassani, että teoksen monimuotoinen kerronta korostaa sekä omaelämäkerran kykyä käsitellä erityisiä ja henkilökohtaisia kokemuksia että sen kykyä kurottaa kohti kulttuurien jaettuja ymmärtämisen ja kokemisen tapoja. Ullin identiteetti kiinnittyy teoksessa 1980-luvun vastakulttuureihin, joita kuvataan dokumentaarisen tarkasti. Subjektiivista kokemusmaailmaa, joka kulkee kerronnassa historiallisen todellisuuden rinnalla, lähestytään puolestaan ekspressiivisin ilmaisukeinon. Teoksen lähtökohtana ovat tekijän henkilökohtaiset kokemukset, mutta kerronta punoo tarinan kulttuurimme yhteisiin kertomusmalleihin ja myytteihin. Omaelämäkerrallinen tarina tarjoaa keinon käsitellä tekijän minuuden lisäksi myös historiallis-sosiaalista todellisuutta ja maailmasta kertomisen tapoja. Tässä piilee omaelämäkerran poliittinen potentiaali, jota myös Lustin teos hyödyntää kommentoidessaan sukupuolten välistä epätasa-arvoa.

Chipless RFID Tag Using Multiple Microstrip Open Stub Resonators

A novel compact RFID tag employing open stubs in a microstrip transmission line is proposed. The prototype of the tag is fabricated on a substrate of dielectric constant 4.4 and loss tangent 0.0018. The tag consists of microstrip open stub resonators and cross polarized transmitting and receiving disc monopole antennas. A prototype of 8 bit data encoded tag is demonstrated in this communication. Method for enhancing the performance of the RFID tag is also proposed. Magnitude or group delay response can be used to decode the tag informations

Frequency Coded Chipless RFID Tag using Spurline Resonators

A novel compact chipless RFID tag using spurline resonators is discussed in this paper. The detection of the tag's ID is using the spectral signature of a spurline resonator circuit. The tag has a data capacity of 8-bits in the range 2.38 to 4.04 GHz. The tag consists of a spurline multiresonating circuit and two cross polarized antennas. The prototype of the tag is fabricated on a substrate CMET/ LK4.3 of dielectric constant 4.3 and loss tangent 0.0018. The measured results show that group delay response can also be used to decode the tag’s identity

5. Krypto-Tag - Workshop über Kryptographie

Dieser Tagungsband enthält die gesammelten Zusammenfassungen der acht eingereichten Vorträge des 5. Krypto-Tags. Der Kryptotag ist eine zentrale Aktivität der Fachgruppe "Angewandte Kryptologie" der Gesellschaft für Informatik e.V. Er ist eine wissenschaftliche Veranstaltung im Bereich der Kryptologie und von der organisatorischen Arbeit der Fachgruppe getrennt.