bluuebunny/crossref_metadata_2025 huggingface dataset:
Linear thermal expansion of the building stone from plutonic massifs of the Vardar zone of Serbia
Authors: Kurešević Lidja , Lužaić Dejan , Delić-Nikolić Ivana
Published: January 2021

Thermal properties of the building stone used for cladding and flooring, especially in exteriors, are significant for durability and lifespan of buildings' surfaces covered with it, but also their aesthetic properties in the sense of integrity. Linear thermal expansion of natural stone units is determined in laboratory examinations, aiming to provide the value of yield between the units as free space for their dilatation and contraction upon temperature oscillations. Where this space is not provided, diurnal changes, especially temperature increase, create stone expansion and tension which can lead to deformations of stone units and mar units' and covered surfaces' integrity and aesthetics. Data on linear thermal expansion values of natural stone, especially from Serbia, are very rare in the literature, so our aim is to contribute to this type of knowledge. Examination of thermal dilatation of stone originating from eight plutonic massifs in External Vardar zone of Serbia has shown that the values are very small, parts of millimetres, and lower than those of the similar rock types originating from China, Brazil and Eastern Serbia. A clear grouping of the values of thermal expansion of the stone according to the genetic groups to which the plutons belong to can be observed. Mineral composition, grain size and presence of metallic minerals in samples have shown no clear influence on thermal dilatation values, but fabric inhomogeneity appears to cause the increase of these values.
Towards innovative and sustainable education in primary schools
Authors: Kovačević Aleksandra , Pendić Zoran
Published: January 2021

One of the most important human rights is the right to quality education for all. It is embedded in the foundations of the UNESCO mission and is contained in the Universal Declaration of Human Rights of 1948 and many other international human rights instruments. It is embedded in the foundations of the UNESCO mission and is contained in the United Nations Universal Declaration of Human Rights of December 10, 1948 and many other international human rights instruments. The United Nations has defined priorities for the development of human civilization until 2030 in the form of 17 goals of sustainable development goals. Education is the fourth element of this set of goals, because it is an indisputable fact that only an educated society is capable of solving problems aimed at achieving all goals. Primary education that children receive in primary schools is especially important, because the success of the entire system of formal and non-formal education in one country largely depends on the quality of this education. It is also the beginning of a marathon of lifelong learning. We must not forget that this education largely depends on the quality of preschool education of children and the involvement of their parents in the process of early stages of education. The paper will include our vision of innovative and sustainable education in primary schools, as well as proposals for continuous improvement of the quality of this education in accordance with the requirements of relevant international standards and accepted world, but also our, experience in primary education.
Measures to reduce pollutant emissions from taxiing aircraft at Belgrade airport
Authors: Mirković Bojana , Ganić Emir
Published: January 2021

This paper presents the analysis o f measures to reduce pollutant emissions o f taxiing aircraft at Belgrade Airport, motivated by one o f the Flighpath 2050 goals that aircraft movements should be emission-free when taxiing. Fuel burn and pollutant emissions are quantified for basic scenario for year 2019, and compared to three alternative scenarios: single engine taxiing, dispatch towing and electric taxiing. For a mid-size airport like Belgrade Airport the priority should be given to single engine taxi procedure because it is not associated with additional implementation costs, unlike the other two scenarios; and estimated benefit is only somewhat smaller compared to electric taxiing case and almost the same as in dispatch towing case.
Switching characteristics of anthraquinone molecular devices based on graphene electrodes
Authors: Cui Yan , Xia Cai-Juan , Su Yao-Heng , Zhang Bo-Qun , Zhang Ting-Ting , Liu Yang , Hu Zhen-Yang , Tang Xiao-Jie ,   School of Science, Xi’an Polytechnic University, Xi’an 710048, China
Published: January 2021

With the development of microelectronics and the miniaturization of electronic devices, the use of molecular materials to construct various components in electronic circuits has become a most likely development trend. Compared with silicon-based semiconductor components, molecular electronic device has the advantages of small size, high integration, low energy consumption and fast response. In recent years, more and more molecules have been used to design molecular devices such as molecular diodes, molecular switches, molecular field effect transistors and molecular memories. In this paper, sandwich structure devices based on graphene nanoribbon electrodes are constructed. The first-principles calculation method combining density functional theory and non-equilibrium Green’s function is adopted to design the molecular devices with functional characteristics. The effects of redox reactions on the electrical transport properties of molecular devices are systematically discussed. The main research contents of this paper are as follows. The switching characteristics of an anthraquinone molecular device based on graphene electrode are studied. The zigzag-edge nanoribbons and armchair-edge graphene nanoribbons are selected as electrodes. Considering the two isomers of anthraquinone (HQ) and anthraquinone (AQ) molecules in the redox reaction, the double electrode molecular junction is constructed. The effects of redox reaction and electrode structure on the switching characteristics of anthraquinone molecular devices are discussed. It is found that the current in the HQ configuration is significantly greater than that in the AQ configuration, regardless of the zigzag-edge graphene electrode or the armchair-edge graphene electrode. That is, under the redox reaction, the anthraquinone molecules show significant switching characteristics. The switching ratio of zigzag-edge graphene electrode is selected to reach a maximum of 3125, and that of armchair-edge graphene electrode is selected to maximum of 1538. In addition, when the armchair-edge graphene is used as an electrode in the HQ configuration, the negative differential resistance is obviously between 0.7 and 0.9 V.
Single event effect and total dose effect of GaN high electron mobility transistor using heavy ions and gamma rays
Authors: Chen Rui , Liang Ya-Nan , Han Jian-Wei , Wang Xuan , Yang Han , Chen Qian , Yuan Run-Jie , Ma Ying-Qi , Shangguan Shi-Peng ,   Center for Space Science and Applied Research, Chinese Academy of Sciences, Beijing 100190, China,   University of Chinese Academy of Sciences, Beijing 100049, China
Published: January 2021

The single event effect (SEE) and the total ionizing dose (TID) effect of a commercial enhancement mode gallium nitride (GaN) high electron nobility transistor (HEMT) with p-type gate structure and cascode structure are studied by using the radiation of heavy ions and 60Co gamma in this paper. The safe operating areas ofSEE, the sensitive parameters degradation of TID effect and the SEE and TID characteristics of GaN HEMT device are respectively presented. The experimental results show that the SEE and TID effect have less influence on the p-type gate GaN device. The linear energy transfer (LET) threshold of the single event Burnout effect (SEB) is higher than 37 MeV·cm2/mg and the failure threshold of TID effect is above 1M rad (Si) for p-type gate GaN device. However, the GaN HEMT device with cascode structure is much more sensitive to SEE and TID effect than p-type gate GaN device. Under heavy ions at LET of 22 MeV·cm2/mg and a cumulative dose of 200 krad (Si), the SEB phenomenon and parameters-degradation of cascode-type GaN HEMT are respectively observed. Besides, the circuit structure of the cascode-type GaN HEMT device is analyzed by using metallographic microscope imaging and focused ions beam technology. It reveals the possible reason why it is sensitive to SEB and TID for cascode-type GaN HEMT. These results show that the extra carriers caused by heavy ion radiation can tunnel the Schottky barrier formed by gate metal and AlGaN layer, leading to a large source-drain current in GaN HEMT device. Meanwhile, it is shown that the metal oxide semiconductor field-effect transistor in cascode circuit for TP90H180PS GaN HEMT may be the main reason why the cascode-type GaN HEMT is sensitive to TID.
Metabolomic profiling of rare cell populations isolated by flow cytometry from tissues
Authors: DeVilbiss Andrew W , Zhao Zhiyu , Martin-Sandoval Misty S , Ubellacker Jessalyn M , Tasdogan Alpaslan , Agathocleous Michalis , Mathews Thomas P , Morrison Sean J
Published: January 2021

Little is known about the metabolic regulation of rare cell populations because most metabolites are hard to detect in small numbers of cells. We previously described a method for metabolomic profiling of flow cytometrically isolated hematopoietic stem cells (HSCs) that detects 60 metabolites in 10,000 cells (Agathocleous et al., 2017). Here we describe a new method involving hydrophilic liquid interaction chromatography and high-sensitivity orbitrap mass spectrometry that detected 160 metabolites in 10,000 HSCs, including many more glycolytic and lipid intermediates. We improved chromatographic separation, increased mass resolution, minimized ion suppression, and eliminated sample drying. Most metabolite levels did not significantly change during cell isolation. Mouse HSCs exhibited increased glycerophospholipids relative to bone marrow cells and methotrexate treatment altered purine biosynthesis. Circulating human melanoma cells were depleted for purine intermediates relative to subcutaneous tumors, suggesting decreased purine synthesis during metastasis. These methods facilitate the routine metabolomic analysis of rare cells from tissues.
Gender-related disparities in the frequencies of PD-1 and PD-L1 positive peripheral blood T and B lymphocytes in patients with alcohol-related liver disease: a single center pilot study
Authors: Kasztelan-Szczerbinska Beata , Adamczyk Katarzyna , Surdacka Agata , Rolinski Jacek , Michalak Agata , Bojarska-Junak Agnieszka , Szczerbinski Mariusz , Cichoz-Lach Halina
Published: January 2021

Background

Exposure to excessive alcohol consumption dysregulates immune signaling. The programed cell death 1 (PD-1) receptor and its ligand PD-L1 play a critical role in the protection against immune-mediated tissue damage. The aim of our study was evaluation of the PD-1/PDL-1 expression on peripheral T and B lymphocytes, its correlation with markers of inflammation and the severity of liver dysfunction in the course of alcohol-related liver disease (ALD).

Material and Methods

Fifty-six inpatients with ALD (38 males, 18 females, aged 49.23 ± 10.66) were prospectively enrolled and assigned to subgroups based on their: (1) gender, (2) severity of liver dysfunction (Child-Pugh, MELD scores, mDF), (3) presence of ALD complications, and followed for 30 days. Twenty-five age- and gender-matched healthy volunteers served as the control group. Flow cytometric analysis of the PD-1/PD-L1 expression on peripheral lymphocyte subsets were performed.

Results

General frequencies of PD-1/PD-L1 positive T and B subsets did not differ between the ALD and control group. When patients were analyzed based on their gender, significantly higher frequencies of PD1/PD-L1 positive B cells in ALD females compared to controls were observed. ALD females presented with significantly higher frequencies of PD-1+ and PD-L1+ B cells, as well as PD-L1+ all T cell subsets in comparison with ALD males. The same gender pattern of the PD-1/PDL1 expression was found in the subgroups with mDF > 32 and MELD > 20. No correlations of PD-1+ and PD-L1+ lymphocyte percentages with mDF, CTP and MELD scores, nor with complications of ALD were observed. Significant correlations of PD-L1 positive B cell frequencies with conventional markers of inflammation were found.

Conclusions

Gender-related differences in the frequencies of PD-1/PD-L1 positive T and B cells were observed in patients with ALD. Upregulation of PD-1+/PD-L1+ lymphocytes paralleled both the severity of alcoholic hepatitis and liver dysfunction in ALD females.
Analytical isoelectric focusing of rat intestinal brush‐border enzymes: postnatal changes and effect of neuraminidase in vitro
Authors: Kraml Jiří , Kolínska Jiřina , Kadlecová Libuše , Zákostelecka Marie , Lojda Zdeněk
Published: July 2002

Increase in p1 values was found between 11 and 30 days of postnatal life in papain‐solubilized brush‐border enzymes of rat small intestine by means of thin‐layer analytical isoelectric focusing in agarose gel. Treatment with neuraminidase converted the acidic forms of enzymes from 11‐ and 20‐day‐old rats into the zymogram patterns identical with the adult, more basic forms. The zymograms of the respective enzymes are the same in 30‐, 60‐ and 74‐day‐old animals and do not change on treatment with neuraminidase.
In vivo modulation of rat liver acyl‐coenzyme A:cholesterol acyltransferase by phosphorylation and substrate supply
Authors: Suckling Keith E. , Stange Eduard F. , Dietschy John M.
Published: July 2002

Rats were infused intravenously with a bolus of mevalonolactone or cholesterol‐rich chylomicron and microsomes prepared from the livers at times up to 2 h after the infusion. Acyl‐CoA cholesterol acyltransferase (ACAT) activity was measured in the microsomes under conditions that define the activity state of the enzyme which recent evidence suggests may depend on the degree of phosphorylation of the enzyme. ACAT activity was increased by both treatments. Some of the observed increase could be attributed to an increase in the supply of substrate cholesterol to the enzyme. The remaining change in activity was consistent with a change in the activity state of ACAT occurring in vivo in response to the influx of cholesterol to the enzyme
A Tris‐induced change in the midpoint potential of Z, the donor to photosystem II, as determined by the kinetics of the back reaction
Authors: Yerkes Christine T. , Babcock Gerald T. , Crofts Antony R.
Published: July 2002

A new method of measuring the rate of the back reaction from the state Z+ P680 QA
− in Tris‐washed chloroplasts is described. By using ratios of back reaction rates we demonstrate a Tris‐induced change in the equilibrium between Z and P680 and attribute this change to an alteration of the midpoint potential of Z by Tris treatment. We also demonstrate that the previously observed inhibition of the back reaction by ADRY reagents can be localized at Z and understood in terms of electron donation to Z+ by ADRY reagents.
The reversible association of quinate:NAD+ oxidoreductase from carrot cells with a putative regulatory subunit depends on light conditions
Authors: Graziana Annick , Ranjeva Raoul , Salimath Bharathi P. , Boudet Alain M.
Published: July 2002

The transfer of carrot cell‐suspension cultures from light to dark conditions provoked considerable and reversible changes in the regulatory and structural properties of quinate:NAD+ oxidoreductase. Thus, the enzyme became directly activatable by Ca2+ and its M
r shifted from 42 kDa to 110 kDa. The analysis of the dark enzyme, purified to homogeneity, showed that an additional subunit of 60–63 kDa was associated to QORase. The homogeneous native enzyme retained the sensitivity to calcium. Preincubation with trypsin had no effect on sensitivity to Ca2+ while a chaotropic agent mimicked the action of Ca2+. It is concluded that light—dark transitions provoke the association of QORase with a putative regulatory subunit which may be a calciprotein.
Separation of Fe+3 from transferrin in endocytosis
Authors: Rao K. , van Renswoude J. , Kempf C. , Klausner R.D.
Published: July 2002

NH4Cl and monensin, two agents which neutralize intracellular acidic compartments, block the segregation of iron from transferrin after endocytosis, while neither of these reagents affects internalization of diferric transferrin into the cell. In conclusion the molecular separation of iron from transferrin inside the cell requires a non‐lysosomal acidic compartment
Cyclic activity of L‐asparaginase through reversible phosphorylation in Leptosphaeria michotii
Authors: Jerebzoff Stéphan , Jerebzoff-Quintin Simonne
Published: July 2002

Asparaginase in L. michotii has previously been shown to have an activity rhythm, the mechanisms of which were investigated. In vitro activation, or reactivation after dephosphorylation, of the partially (200‐fold) purified asparaginase with protein kinase activity was obtained by ATP or Pi addition; these effects varied according to the phase of the activity rhythm at which enzyme was extracted. A high‐M
r aggregate with asparaginase activity was phosphorylated by [γ‐32P]ATP. By SDS‐electrophoresis of dephosphorylated asparaginase a ∼60‐kDa 32P‐labelled protein with alkaline phosphatase activity became detectable. Regulation of the asparaginase activity rhythm in L. michotii is dependent on a reversible phosphorylation process.
The complete amino acid sequence of the ribosomal protein HS3 from Halobacterium marismortui, an archaebacterium
Authors: Kimura Makoto , Langner Gabriele
Published: July 2002

The complete amino acid sequence of the ribosomal protein HS3 from Halobacterium marismortui has been determined, which is, to our knowledge, the first complete protein sequence from an archaebacterial ribosome. Comparison of the primary structure of HS3 with all sequenced ribosomal proteins from other organisms revealed that HS3 is significantly homologous with protein S9 from the gram‐negative Escherichia coli and the gram‐positive Bacillus stearothermophilus, having 27 and 37% identical residues, respectively. Based on these and other results it is possible that some ribosomal components of archaebacteria are related to eubacteria, especially gram‐positive bacteria, and others to eukaryotes.
Reconstitution of Azotobacter vinelandii ferredoxin I as a {2[4Fe‐4S]1+/2+} protein
Authors: Morgan T.V. , Stephens P.J. , Burgess B.K. , Stout C.D.
Published: July 2002

As normally isolated, Azotobacter vinelandii ferredoxin I (Fd I) is a {[4Fe‐4S],[3Fe‐3S]} protein: (7Fe)Fd I. We report that anaerobic reconstitution of Fd I from its apoprotein yields a protein whose spectra are distinct from those of (7Fe)Fd I and typical of bacterial ferredoxins. We identify this new form of Fd I as a {2[Fe‐4S]} protein: (8Fe)Fd I. (8Fe)Fd I is unstable in air and decomposes to give a ∼ 10% yield of (7Fe)Fd I. These results increase the probability that (8Fe)Fd I is the form of Fd I occurring in vivo and that (7Fe)Fd I results from oxidative degradation during purification.
The L7/L12 proteins change their conformation upon interaction of EF‐G with ribosomes
Authors: Gudkov A.T. , Gongadze G.M.
Published: July 2002

The different functional complexes of ribosomes with elongation factor F (EF‐G) were studied by digestion experiments with trypsin. It was found that upon interaction of EF‐G with ribosomes the L7/L12 proteins are sensitive to trypsin and are trypsin resistant after dissociation of EF‐G from ribosomes. The significance of conformational alterations in the L7/L12 and also in the other proteins in the translation process is discussed.
Intracellular pH measurements using the fluorescence of 9‐aminoacridine
Authors: Vigne Paul , Frelin Christian , Lazdunski Michel
Published: July 2002

9‐Aminoacridine is a fluorescent weak base which distributes across the plasma membrane according to the pH gradient, and can be used to monitor changes in intracellular pH in eukaryotic cells. A calibration technique using the ionophore nigericin is described which allows us to measure intracellular pH values of chick skeletal muscle cells.
Reconstitution of ornithine transport in liposomes with Lubrol extracts of mitochondria
Authors: Hommes F.A. , Eller A.G. , Evans B.A. , Carter A.L.
Published: July 2002

The ornithine translocase of beef liver mithochondria was extracted with Lubrol WX and reconstituted in liposomes. The uptake of ornithine by the reconstituted vesicles followed Michaelis‐Menten kinetics, and was dependent on the intraliposomal pH, the time of sonication of the reconstituted liposomes and the phospholipid to detergent ratio. It is concluded that the ornithine translocator can be reconstituted, which makes the purification of this translocator feasible.
Erythrocyte membrane acyl : CoA synthetase activity
Authors: Davidson B.C. , Cantrill R.C.
Published: July 2002

The presence of long‐chain acyl : CoA synthetases in mammalian microsomes and mitochondria has been established previously [(1971) Biochim. Biophys. Acta 231, 32‐47]. The presence of a plasma membrane‐associated enzyme was investigated in human erythrocyte ghost plasma membranes, where an enzyme exhibiting high activity, and with a preferred substrate of 18 carbon chain length, was discovered. The results are consistent with the presence of a single enzyme. The effect of the degree of Unsaturation of the fatty acid substrates was not as pronounced as that arising from the length of the carbon chain. The pattern of substrate preference of the enzyme was ω3 polyenoics > ω6 polyenoics > ω9 monoenoics > saturated fatty acids. This may relate to the similar substrate preference pattern exhibited by the fatty acyl desaturase enzymes. However, the role played by long‐chain acyl: CoA synthetase in erythrocyte metabolism is uncertain, but may relate to the transportation of polyenoic fatty acids in the circulation.
A phospholipid transfer protein that binds long‐chain fatty acids
Authors: Rickers Jutta , Spener Friedrich , Kader Jean-Claude
Published: July 2002

A phospholipid transfer protein, purified from spinach leaves, is able to bind long‐chain fatty acids (oleic and linoleic acids) as well as oleoyl‐CoA. This binding was demonstrated by analytical isoelectric focusing and electrophoretic titrations. The finding that a protein, known for its ability to catalyze intermembrane transfers of phospholipids, is also able to bind acyl chains, opens new perspectives for the physiological significance of this category of proteins in plant cells.
500 MHz 1H‐NMR study of the interaction of daunomycin with B and Z helices of d(CGm5CGCG)
Authors: Neumann J.M. , Cavailles J.A. , Herve M. , Tran-Dinh S. , d'Estaintot B.Langlois , Huynh-Dinh T. , Igolen J.
Published: July 2002

The interaction of daunomycin with B and Z helices of a self‐complementary DNA fragment d(CGm5CGCG) in solution was studied by 1H‐NMR spectroscopy at 500 MHz. The results show that the B‐Z transition kinetics is not affected by addition of daunomycin. Daunomycin binds exclusively to the B form of d(CGm5CGCG). Z exchanges with B while the latter also exchanges with the B duplex‐daunomycin complexes.
Inhibition by forskolin of cytosolic calcium rise, shape change and aggregation in quin2‐loaded human platelets
Authors: Sage S.O. , Rink T.J.
Published: July 2002

The adenylate cyclase stimulator forskolin was used to study the inhibitory effects of elevated cAMP on the activation of washed human platelets loaded with the fluorescent Ca2+ indicator quinl. In the presence of 10 μM isobutylmethylxanthine forskolin inhibited rises in [Ca2+]i evoked by thrombin and platelet‐activating factor (PAF) due to both Ca2+ influx and release from internal stores with similar potency. Aggregation evoked by thrombin and PAF was suppressed whilst partial shape‐change persisted, even in the absence of a measurable rise in [Ca2+]i. Forskolin did not affect the rise in [Ca2+
i evoked by Ca2+ ionophore; aggregation was suppressed but shape‐change persisted.
On the mechanism of enzymatic lignin breakdown
Authors: Schoemaker H.E. , Harvey P.J. , Bowen R.M. , Palmer J.M.
Published: July 2002

The recent discovery of an extracellular enzyme from Phanerochaete chrysosporium capable of degrading lignin model compounds has countered the view that reactive diffusible oxygen species are responsible for lignin biodegradation. In this paper we propose a mechanism which accounts for both the results obtained in enzymatic lignin degradation studies and in studies involving active oxygen species. The quintessence of the mechanism is initial one‐electron oxidation of the lignin model compounds or of a specific lignin subunit followed by subsequent breakdown reactions via radical cation intermediates. The implication of this type of mechanism on the oxidative biodegradation of the natural lignin polymer is discussed.
Generation of the electrochemical potential of Na+ by the Na+‐motive NADH oxidase in inverted membrane vesicles of Vibrio alginolyticus
Authors: Tokuda Hajime , Udagawa Toshiaki , Unemoto Tsutomu
Published: July 2002

Inverted vesicle
Na+ pump
NADH oxidase
Marine bacterium
Na+ electrochemical potential
Respiration
Chemical‐enzymatic incorporation of D‐amino acids into peptides: synthesis of diastereomeric (D‐Ala2, D‐Leu5)enkephalinamides
Authors: Stoineva I.B. , Petkov D.D.
Published: July 2002

Enzymatic synthesis
Opioid peptide
Enkephalin
NMR
Aromatic ring cleavage of veratryl alcohol by Phanerochaete chrysosporium
Authors: Leisola Matti S.A. , Schmidt Brigitte , Thanei-Wyss Ursula , Fiechter Armin
Published: July 2002

Two oxidation products of veratryl alcohol were isolated from ligninolytic cultures of Phanerochaete chrysosporium. After purification the compounds were characterized by 1H‐NMR, mass spectrometry and infrared spectrometry. The structural information suggests that the compounds are two isomers of a ring cleavage product from veratryl alcohol. Both compounds were also found as by‐products when veratryl alcohol was oxidized to veratraldehyde with the crude extracellular ligninase preparation.
Release of T‐kinin and bradykinin in carrageenin induced inflammation in the rat
Authors: Barlas Aydin , Sugio Kazuo , Greenbaum Lowell M.
Published: July 2002

T‐kinin
T‐kininogen
Bradykinin
Inflammation
Acute‐phase protein
Carrageenin
Morphological properties of proteoliposomes reconstituted with the Na+ pump methylmalonyl‐CoA decarboxylase from Veillonella alcalescens
Authors: Rohde Manfred , Däkena Patricia , Mayer Frank , Dimroth Peter
Published: July 2002

Proteoliposomes reconstituted with purified methylmalonyl‐CoA decarboxylase have a mean diameter of 62 nm and contain on average 9–10 enzyme molecules integrated in their membrane. Affinity labelling with avidin‐gold complexes was used to determine the number of enzyme molecules which were oriented with the biotin prosthetic group exposed to the outside. About 50% of the proteoliposomes became labelled under optimized conditions, containing on average 3.4 avidin‐gold complexes on their surface. These data from electron microscopy are in good agreement with those from biochemical analyses indicating the presence of about 7 enzyme molecules per average‐sized proteoliposome from which about 1–2 enzyme molecules are properly oriented in the membrane to perform an inwardly directed Na+ pumping.
Expression of the cloned subunits of Escherichia coli transhydrogenase from separate replicons
Authors: Clarke David M. , Bragg Philip D.
Published: July 2002

The pntA and pntB genes of Escherichia coli, encoding the α‐ and β‐subunits of the pyridine nucleotide transhydrogenase, were cloned individually in two different compatible plasmids into Escherichia coli mutants lacking transhydrogenase activity. Energy‐linked and non‐energy‐linked transhydrogenase activities were produced only in cells carrying both plasmids thus showing that the products of both genes are required for the formation of an active enzyme. ATP‐energized transhydrogenase activity was not increased in cells containing amplified levels of the transhydrogenase when the cell membrane ATPase was also amplified. It is suggested that the excess transhydrogenase is effectively uncoupled from the ATPase by compartmentalization in the cell.
Modulation of K+ conductance by intracellular pH in pancreatic β‐cells
Authors: Rosario Luis M. , Rojas Eduardo
Published: July 2002

Measurements of the effects of NH3/NH4
+ on glucose‐induced electrical activity in β‐cells from microdis‐sected mouse islets of Langerhans and on intracellular pH in single collagenase‐isolated islets pre‐loaded with a fluorescent pH probe were performed and are reported here. Application of NH3/NH+
4 (15 mM) in the presence of glucose (11 mM) promptly hyperpolarized the β‐cell membrane, reduced input resistance by 60% and blocked electrical activity. These changes were paralleled by an increase in islet fluorescence indicative of a cytosolic pH increase. Removal of NH4Cl initially stumulated electical activity, which returned to resting level with a time constant of 51 s. Concomitant with the removal of NH4C1 there was a drop in pHi followed by a slow return to resting level with a time constant of 83 s. The results suggest that the [Ca2+]‐dependent K+ channel in the β‐cell membrane is activated by a rise in cytosolic pH.
Chemical cross‐linking of ferredoxin to spinach thylakoids
Authors: Merati Giuliana , Zanetti Giuliana
Published: July 2002

Ferredoxin has been chemically cross‐linked to thylakoids by using N‐ethyl‐3‐(3‐dimethylaminopropyl)‐carbodiimide. The membranes thus treated became able to photoreduce cytochrome c and to catalyze the NADPH‐cytochrome c reductase reaction without adding exogenous ferredoxin. Preincubation of thylakoids with an antibody against ferredoxin‐NADP+ reductase before carbodiimide treatment or removal of the reductase by mild trypsin treatment after the cross‐linking reaction did not alter the cytochrome c photoreduction activity of the treated membranes. Two independent binding sites of ferredoxin to thylakoids are thus inferred: one site is shown to be the membrane‐bound reductase, the second is suggested to be at the level of the photosystem I complex.
Primary structure of the acidic carbohydrate chain of hemocyanin from Panulirus interruptus
Authors: van Kuik J.A. , Breg J. , Kolsteeg C.E.M. , Kamerling J.P. , Vliegenthart J.F.G.
Published: July 2002

The N‐glycosidic carbohydrate chains of hemocyanin from the spiny lobster Panulirus interruptus were liberated by hydrazinolysis of a pronase digest and subsequently reduced. Separation of the mixture of oligosaccharide‐alditols by high‐voltage paper electrophoresis resulted in a neutral (90%) and an acidic (10%) fraction. 500‐MHz 1H‐NMR spectroscopy of the acidic fraction revealed a single component with the following novel structure:
Effects of protein kinase C activation on human platelet cyclic AMP metabolism
Authors: Bushfield Mark , Hopple Sara L. , Gibson Iain F. , Murdoch Fiona A. , MacIntyre D.Euan
Published: July 2002

Treatment of intact human platelets with the tumour‐promoting phorbol ester, phorbol 12‐myristate 13‐acetate (PMA), specifically inhibited PGD2‐induced cyclic AMP formation without affecting the regulation of cyclic AMP metabolism by PGI2, PGE1, 6‐keto‐PGE1, adenosine or adrenaline. This action of PMA was: (i) concentration‐dependent; (ii) not mediated by evoked formation or release of endogenous regulators of adenylate cyclase activity (thromboxane A2 or ADP); (iii) mimicked by 1,2‐dioctanoylglycerol (DiC8) but not by 4α‐phorbol 12,13‐didecanoate (which does not activate protein kinase C); (iv) attenuated by Staurosporine. These results indicate that activation of protein kinase C in platelets may provide a regulatory mechanism to abrogate the effects of the endogenous adenylate cyclase stimulant PGD2 without compromising the effects of exogenous stimulants of adenylate cyclase (PGI2, 6‐keto‐PGE1, adenosine).
Stoichiometry and turnover of photosystem II polypeptides
Authors: Gounaris Kleoniki , Pick Uri , Barber James
Published: July 2002

Using Triton X‐100 solubilization and sucrose gradient fractionation, a photosystem II core complex has been isolated from the green alga Dunaliella salina. The complex was similar to that previously isolated from spinach [(1985) FEBS Lett. 188, 68‐72] being composed of five main polypeptides of apparent molecular masses, 47, 40, 32, 30 and 10 kDa. Studies using 14C indicate that with either dark or illuminated cells the Stoichiometry of these polypeptides is 1:1:1:1:1 with about 20 chlorophyll a molecules per monomeric unit. According to 35S labelling only the 32 kDa polypeptide, identified as the D1 or QB binding protein by immunoblotting, is rapidly turning over at a rate greater than the growth rate. The level of labelled sulphur associated with each polypeptide is reasonably consistent with the amino acid content derived from gene sequences and identifies the 47, 40, 32, 30 and 10 kDa apoproteins as products of the psbB, psbC, psbA, psbD and psbE genes.
Evidence for the presence of a cytosine deaminase that does not catalyze the deimination of creatine
Authors: Kim Jong Min , Shimizu Sakayu , Yamada Hideaki
Published: July 2002

The presence of a cytosine deaminase, which shows no activity on creatinine, was first noted in Alcaligenes denitrificans ssp. denitrificans J9 and Arthrobacter sp. J11. DEAE‐Sephacel column chromatography of cellfree extracts of these microorganisms gave a single peak for cytosine deaminase activity, and no fraction contained creatinine deiminase activity. The DEAE‐Sephacel active fractions from A. deinitrificans ssp. denitrificans J9 were resolved into two fractions showing cytosine deaminase activity upon Sephacryl S‐200 column chromatography; the larger enzyme was termed cytosine deaminase I and the smaller one cytosine deaminase II. Both cytosine deaminases did not deaminate creatinine but did deaminate 5‐fluorocytosine rapidly. The molecular masses of cytosine deaminases I and II were estimated to be 200 and 37 kDa, respectively.
Secretion of human insulin by a transformed yeast cell
Authors: Thim Lars , Hansen Mogens T. , Sørensen Anders R.
Published: July 2002

A yeast expression plasmid encoding a mini‐proinsulin molecule was constructed and transformed into Saccharomyces cerevisiae. The plasmid encoded the sequence: B‐Arg‐Arg‐Leu‐Gln‐Lys‐Arg‐A in which B represents the B‐chain (30 amino acid residues) and A represents the A‐chain (21 amino acid residues) of human insulin. The secreted peptides were shown to be a mixture of human insulin and des(B‐30)human insulin. Thus, correct disulphide bridges can be established in proinsulin‐like molecules devoid of a normal C‐peptide region. Furthermore, the specificity of the yeast processing enzymes is so similar to the proinsulin converting enzymes in the human pancreatic β‐cell that it allows the processing of the mini‐proinsulin to insulin.
Differential splicing in the 3′ non‐coding region of rat cytochrome P‐452 (P450 IVA1) mRNA
Authors: Earnshaw David , Dale Jeremy W. , Goldfarb Peter S. , Gibson G.Gordon
Published: July 2002

The gene coding for the clofibrate‐induced cytochrome P‐452 has been isolated from a cDNA library and a full‐length cDNA clone sequenced. The sequence was identical with that reported by Hardwick et al. [(1987) J. Biol. Chem. 262, 801–810] apart from the presence of an additional 75 nucleotides inserted adjacent to the 3′‐end of the coding region. Northern blot analysis indicates that this is probably a consequence of differential splicing, which may be relevant to the tissue‐specific regulation of this gene.
Inhibition and labelling of the mitochondrial 2‐oxoglutarate carrier by eosin‐5‐maleimide
Authors: Zara V. , Palmieri F.
Published: July 2002

Unlike hydrophobic maleimides, eosin‐5‐maleimide and to a lesser extent other relatively polar maleimides inhibit the 2‐oxoglutarate carrier of bovine heart mitochondria. The impermeable eosin‐5‐maleimide labels the 2‐oxoglutarate carrier in intact mitochondria but not in submitochondrial particles. 2‐Oxoglutarate protects the carrier against inactivation by eosin‐5‐maleimide and decreases the fluorescence associated with the purified protein. Other anions which are not substrates of the carrier have no protective effect. It is concluded that sulfhydryl groups essential for the activity of the 2‐oxoglutarate carrier are located at the cytosolic face of the inner mitochondrial membrane. They appear to be present at the substrate‐binding site and located in a hydrophilic environment.
Expression of the glyceraldehyde‐3‐phosphate dehydrogenase gene from the extremely thermophilic archaebacterium Methanothermus fervidus in E. coli
Authors: Fabry Stefan , Lehmacher Anselm , Bode Wolfram , Hensel Reinhard
Published: July 2002

The gene of glyceraldehyde‐3‐phosphate dehydrogenase (GAPDH) from the extremely thermophilic archaebacterium Methanothermus fervidus (growth optimum 82°C) was cloned in vector pJF118EH and expressed in E. coli cells. As shown by molecular mass determination, protein sequencing, heat stability, and substrate saturation kinetics, the enzyme synthesized in E. coli is identical to the original enzyme from M. fervidus. The high thermostability of the E. coli‐produced M. fervidus GAPDH allows rapid purification to homogeneity. From this enzyme protein crystals were grown which proved to be suitable for X‐ray analysis. The crystals are of tetragonal space group P4122 and contain a dimer per asymmetric unit.
The complex of mitochondrial F1‐ATPase with the natural inhibitor protein is unable to catalyze single‐site ATP hydrolysis
Authors: Kalashnikova T.Yu. , Milgrom Ya.M. , Postanogova N.V.
Published: July 2002

Interaction of mitochondrial F1‐ATPase with the isolated natural inhibitor protein resulting in the inhibition of multi‐site ATP hydrolysis is accompanied by the loss of activity at low ATP concentrations when single‐site hydrolysis should occur. Catalytic site occupancy by [14C]nucleotides in F1‐ATPase during steady‐state [14C]ATP hydrolysis, which is saturated in parallel with single‐site catalysis, is prevented after blocking the enzyme with the inhibitor protein.
Boar proacrosin is a single‐chain molecule which has the N‐terminus of the acrosin A‐chain (light chain)
Authors: Čechová D. , Töpfer-Petersen E. , Henschen A.
Published: July 2002

Boar proacrosin was isolated from spermatozoa by a novel procedure under conditions preventing proenzyme activation. The spermatozoal extract was fractionated by gel filtration and reversed‐phase FPLC, all in acidic solutions. Isolated proacrosin had a molecular mass of 55/53 kDa (doublet) and was devoid of amidolytic activity. Its single N‐terminal sequence corresponded to that of the 23‐residue acrosin A‐chain and continued with that of the acrosin B‐chain. Autoactivation at pH 7.8 did not influence the molecular mass. However, activated material contained two parallel N‐terminal sequences, those of the A‐ and B‐chain. Thus, activation of proacrosin is analogous to that of other serine proteinase proenzymes.
Purification and characterization of phospholipase A2 inhibitory proteins from pig thyroid gland
Authors: Antonicelli F. , Rothhut B. , Martiny L. , Aguie-Aguie G. , Lambert B. , Bellon G. , Russo-Marie F. , Jacquemin C. , Haye B.
Published: July 2002

A 32 kDa phospholipase A2 inhibitory protein was isolated from pig thyroid gland after calcium precipitation and fast protein liquid anion‐exchange chromatography. SDS‐polyacrylamide gel electrophoresis revealed the purity of the protein. The protein activity was assessed by the inhibition of pancreatic phospholipase A2 on [3H]oleic acid‐labelled Escherichia coli membranes as substrate and on the prostaglandin E2 production of cultured thyroid cells. The amino acid composition and the isoelectric point were quite similar to those of endonexin previously described in other tissues or cells. The cross‐reactivity of a polyclonal antibody against a 32 kDa lipocortin from human peripheral blood mononuclear cells with our thyroidal 32 kDa protein confirmed its lipocortin nature. Before the purification by fast protein liquid chromatography, the Ca2+ pellet contained lipocortin I (35 kDa and its core protein 33 kDa) identified by its cross‐reactivity with a polyclonal antibody.
Identification of the inhibitor of the plasminogen activator as the major protein secreted by endothelial rat liver cells
Authors: Kuiper Johan , Kamps Jan A.A.M. , Van Berkel Theo J.C.
Published: July 2002

Freshly isolated Kupffer and endothelial liver cells exhibit a rate of ‘de novo’ protein synthesis which is twice as high per mg cell protein as that of parenchymal liver cells and contribute significantly (7.5% and 5.9%, respectively) to total liver protein secretion. In parenchymal cells the main secretory protein is a 68 kDa protein (containing 19% of the secreted radioactivity, presumably albumin). In Kupffer cells a 49 kDa protein contains 8% of the secreted radioactivity, while in endothelial liver cells a 55 kDa protein is the most prominent secretory protein (containing 11% of the secreted radioactivity). By aid of a specific antibody the 55 kDa protein was identified as the inhibitor of the plasminogen activator and in the liver this protein was only secreted by the endothelial cells.
Signal transduction in EJ‐H‐ras‐transformed cells: De novo synthesis of diacylglycerol and subversion of agonist‐stimulated inositol lipid metabolism
Authors: Chiarugi Vincenzo P. , Magnelli Lucia , Pasquali Franca , Basi Gabriele , Ruggiero Marco
Published: July 2002

We examined the level of 1,2‐diacylglycerol and inositol phosphates in normal and EJ‐H‐ras‐transformed BALB/3T3 fibroblasts by prelabelling the cells with [3H]glycerol, [3H]inositol, [14C]glucose, [14C]arachidonic acid, and [14]palmitic acid. Steady‐state level of diacylglycerol was increased in ras‐transformed cells prelabelled with radioactive glycerol, glucose and palmitic acid. Steady‐state level of inositol phosphates, however, was the same in control and transformed cells. Diacylglycerol labelling by [14C]arachidonic acid was the same in control and transformed cells. Insulin dramatically increased diacylglycerol labelling by [14C]glucose in normal cells, wheras it did not affect ras‐transformed fibroblasts. Neurotransmitter‐induced inositol lipid turnover was greatly enhanced in ras‐transformed cells; conversely, platelet‐derived growth factor and thrombin‐stimulated normal cells to a greater extent than transformed fibroblasts. Taken together these results suggest that ras transformation may induce multifarious effects on signal transduction: it may cause de novo synthesis of diacylglycerol and subversion of neurotransmitter and growth factor receptor coupling to inositol lipid metabolism.
Gene structure of rat cathepsin H
Authors: Ishidoh Kazumi , Kominami Eiki , Katunuma Nobuhiko , Suzuki Koichi
Published: July 2002

The gene structure of rat cathepsin H was determined. It comprises at least 12 exons of various lengths (32–433 bp) spanning in total more than 17.5 kbp. The gene structure does not correspond well to the functional unit of the proteinase. The region around the active site Cys residue, the most conserved region among cysteine proteinases, is split by an intron. This is a common characteristic among the gene structures of cysteine proteinases.
Enhanced levels of cathepsin B mRNA in murine tumors
Authors: Moin Kamiar , Rozhin Jurij , McKernan Timothy B. , Sanders Virginia J. , Fong Dunne , Honn Kenneth V. , Sloane Bonnie F.
Published: July 2002

Relative amounts of mRNA for cathepsin B were measured in normal murine liver and three murine tumors, an invasive liver tumor (hepatoma, Hepa cl 9) and two melanoma variants (B16‐F1 and B16 amelanotic melanoma, B16a). Using a human cDNA to the cathepsin B coding region as a hybridization probe, we detected two species of cathepsin B specific RNA transcripts (2.2 and 4.1 kb) in total RNA preparation of all four tissues. The concentrations of the 2.2 and 4.1 kb species were 3.6 and 2.7‐fold greater in the highly metastatic B16a melanoma than in normal liver. The concentration of the 2.2 kb species in the invasive hepatoma wa 1.7‐fold greater than in normal liver. The increased levels of the 2.2 kb message were reflected in increases in activity of cathepsin B in both Hepa cl 9 and B16a.
Deglycosylation of the chymotryptic collagen‐binding fragment of human plasma fibronectin does not modify its affinity to denatured collagen
Authors: Delannoy Philippe , Montreuil Jean
Published: July 2002

N‐Glycanase deglycosylation of purified 44 kDa chymotryptic collagen‐binding domain from human plasma fibronectin does not significantly modify its behavior on gelatin affinity chromatography. This indicates that carbohydrates do not play any role in the binding affinity of fibronectin to collagen. The influence of changes in glycosylation on the biological functions of fibronectin is discussed.
Influence of non‐esterified fatty acids on respiratory control of reconstituted cytochrome‐c oxidase
Authors: Thiel Carsten , Kadenbach Bernhard
Published: July 2002

Bovine heart cytochrome‐c oxidase was reconstituted in liposomes (asolectin) and the activity measured in the presence and absence of uncoupler at increasing concentrations of non‐esterified fatty acids. Palmitic and stearic acids resulted in a decrease of about 40% in the respiratory control ratio at a concentration of 1 νM, when measured using a spectrophotometric procedure but not with a polarographic assay method. At higher fatty acid concentrations no further change was found. A 50% decrease in respiratory control was determined when the enzyme was reconstituted in pure phosphatidylcholine containing 2% cardiolipin. The respiratory control of reconstituted cytochrome‐c oxidase from bovine liver was not influenced by fatty acids.
Insulin stimulates the translocation of protein kinase C in rat adipocytes
Authors: Ishizuka Tatsuo , Cooper Denise R. , Farese Robert V.
Published: July 2002

Insulin‐induced changes in protein kinase C were examined in cytosol and membrane fractions of rat adipocytes enzymatically after Mono Q column chromatography and by immunoblotting. During a 5–20 min period of insulin treatment, cytosolic protein kinase C decreased by approximately 50%, whereas membrane protein kinase C increased nearly 2‐fold. These findings suggest that insulin stimulates the translocation of protein kinase C in rat adipocytes.
Regulation of glutathione S‐transferase subunits 3 and 4 in cultured rat hepatocytes
Authors: Morel Fabrice , Vandenberghe Yves , Pemble Sally , Taylor John B. , Ratanasavanh Damrong , Rogiers Vera , Ketterer Brian , Guillouzo André
Published: July 2002

mRNA levels of glutathione S‐transferase (GST) subunits 3 and 4 were measured with a specific cDNA probe in adult rat hepatocytes maintained either in conventional culture or in coculture with rat liver epithelial cells. Four media conditions were used, i.e. with or without fetal calf serum (FCS) and with nicotinamide or dimethylsulfoxide (DMSO). When FCS was present in the culture medium, GST subunit 3 and 4 mRNAs were expressed at a level close to that found in freshly isolated hepatocytes during the whole culture period both in conventional culture and in coculture. All other culture conditions resulted in an increase of GST 3 and 4 mRNA levels. After exposure to phenobarbital an increase in GST 3 and 4 mRNA levels was demonstrated in both culture systems. Comparison with previous findings on the expression of GST subunits 1, 2 and 7 in the same culture conditions indicates that the different classes of GST are regulated independently.
Consensus peptide antibodies reveal a widespread occurrence of Ca2+/lipid‐binding proteins of the annexin family
Authors: Gerke Volker
Published: July 2002

Antibodies generated against synthetic peptides that correspond to highly conserved sequence motifs in the annexins reacted with a variety of annexins from different species. These include Xenopus laevis and Drosophila melanogaster, which contain cross‐reacting polypeptides of apparent M
r, 34000 and 30000. As expected for typical annexins, the two Drosophila proteins interact in a Ca2+‐dependent manner with phosphatidylserine liposomes.