bluuebunny/crossref_metadata_2025 huggingface dataset:
Study of Factors Influencing on the Brand Availability at Stores in Sri Lankan Cables Firm
Authors: Sellar Treshalin
Published: May 2021

In the view of the availability of the selected brand cables at the stores, it is a vital aspect that the decision-makers will have to consider that the product and services of the internal aspects will have to synchronize with the external aspects in order to gain the advantage of the brand, competition as well as the marketing channels. Prime objective of the research is to identify and understand the factors that affect the availability of the selected cables at the stores in Sri Lanka in absolute and relative terms in order to provide effective recommendations to increase the product reachability. The research has been considered at all the stores as well as the distributor who sell the cable products all around the selected region. The participants were taken from the Colombo district, and the sample has been considered as a hundred. The study has been designed in a quantitative manner where the research instrument has taken to collect the data was a structured questionnaire. The research has attained the outcomes as there is a relationship among competitor cable products, brand awareness, intermediaries’ activities with the dependent variable of availability of the selected brand cables at the stores. The researcher has figured out the aspects which could motivate the consumers to make the purchasing decisions with regards to the selected variables of the cable products.
Direct and Residual Effect of Integrated Nitrogen Management on Physico-Chemical Properties of Soil
Authors: Deekshitha D. K. D. , Ch Sujani Rao, , Subbaiah P. V. , Luther M. Martin , Rao V. Srinivasa
Published: May 2021

Aim: To find out the direct and residual effect of integrated nitrogen management on physico-chemical properties of soil under rice – maize cropping system.

Study Design: The experiment was laid out in randomized block design during kharif season and in split plot design during rabi crop.

Place and Duration of Study: At Agricultural college farm, Bapatla during 2018-19 and 2019-20 in Kharif and rabi seasons.

Methodology: After the preliminary layout, paddy was sown in kharif and maize was sown in rabi season. Vermicompost and green leaf manure were applied two weeks before sowing of paddy. Treatments applied for paddy were considered as main plots and each one divided into two sub plots during rabi.

Results: Applications of different treatments did not show any significant difference in physico-chemical properties of soil (pH and EC) at all the growth stages of rice and maize during both the years of study. Application of 75% RDN through green leaf manure + 25% RDN through inorganics (T5) to rice during kharif significantly improved organic carbon in both kharif and rabi seasons whereas different levels of fertilizers applied have not shown an significant influence on soil organic carbon content.

Conclusion: Application of inorganic fertilizers, organic manures and their combination didn’t show marked difference on physico-chemical properties of soil like pH and EC. However substitution of 75% N through GLM in kharif season significantly improved OC content in soil during both kharif and rabi and organic carbon content was not influenced by the level of fertilizers from 50% RDF (S1) to100% RDF (S2).
Antibiotic Resistance Profile and Resistance Determination of Bacteria Isolated from Water in Southern Benin
Authors: Koudokpon Hornel , Dougnon Victorien , Lougbegnon Christelle , Deguenon Esther , Mousse Wassiyath , Agbankpe Alidehou Jerrold , Avodagbe Gérardine , Saidou Souad , Bankole Honoré , Baba-Moussa Lamine
Published: May 2021

Background: The environment plays an important role in the dissemination of multidrug resistant bacteria, especially through the aquatic ecosystem, including hospital effluents, rivers, but also spring water and drinking water. This study aims to determine selected antimicrobial resistance genes in some aquatic matrices in southern Benin.

Methods: Collected water samples were filtered through a membrane 0.22 µm thick. After filtration, the membrane was deposited on Muëller Hinton agar. Then the colonies resulting from this subculture were subjected to a microbiological examination by the conventional method. The antibiotic sensitivity test was carried out by the Kirby Bauer method according to the recommendations of the French Society of Microbiology. Resistance genes were looked for by PCR.

Results: Of the 222 water samples collected, 265 bacterial strains were isolated, the majority of which were strains of Coagulase Negative Staphylococcus (CNS) with 37.74% (n = 100), followed by strains of Klebsiella pneumoniae (21.89%; n = 58), Escherichia coli (10.57%; n = 28). All isolated gram-negative bacilli strains are multidrug resistant with resistance of all strains to amoxicillin, ampicillin and amoxicillin + clavulanic acid. Of the 15 resistance genes searched in the genome of Gram-negative bacilli strains, 8 were detected, namely the TEM, SHV, CTX-M15, VIM, NDM, SUL1, SUL2 and AADA genes. Resistance of CNS strains to amoxicillin, oxacillin and cefoxitin was observed. The meca gene was detected in all CNS strains. The vanA and VanB genes were only detected in strains isolated from drinking water in sachets collected from producers and street sellers.

Conclusion: These results show the dissemination of resistance genes in Benin and once again confirms the urgency of a global fight against antimicrobial resistance.
Olfactory Function Assessment: Standardization of a New Quantitative Technique for the Indian Population
Authors: Jacob Navya Jith , Rajkumar D. , Sudha M. , Varghese Varsha , Mukkadan J. K.
Published: May 2021

Aim: Olfactory function assessment is often neglected in clinical settings due to a lack of appropriate cost effective techniques. We therefore aimed to develop a cost effective, reliable and culturally appropriate tool for olfactory function assessment among the Indian population and to compare olfactory functions among 63 healthy controls and 32 idiopathic Parkinson’s disease patients.

Materials and Methods: Olfactory stimuli were applied to the nostrils of the participants using an olfactometer. Five different odoriferous substances common to Indian culture were used for the study in three different concentrations: ginger (1%, 2%, 3%), cardamom (0.4%, 2%, 3%), garlic (0.8%, 1.4%, 2%), coffee (1.6%, 2%, 4%), vanilla (2%, 3%, 4%). Olfactory recognition threshold, olfactory identification score and olfactory discrimination score were observed among the control population and Parkinson’s disease population.

Results: The olfactory recognition threshold was significantly high among the Parkinson’s disease group compared to controls (Mann Whitney U test, p<0.001). Reliability was tested using the test-retest method among the control group and all olfactory variables in three different concentrations had either r value closer to 1 or 1, which shows an acceptable level of reliability. The correlation was found to be significant (p<0.001). A receiver operating characteristic (ROC) curve drawn for olfactory recognition thresholds at different concentrations for the five odouriferous substances and the area was determined to classify cases and controls (Determined areas: ginger = 0.928, cardamom = 0.955, garlic = 0.921, Coffee = 0.950, vanilla = 0.950). The area under the curve was found to be significant in classifying the cases and the control.

Conclusion: The newly developed olfactory assessment tool was found to be reliable and effective in assessing olfactory parameters like recognition threshold, identification score and discrimination score among the Indian population.
The Frozen Section Yesterday and Today: Pediatric Solid Tumors—Crucial Issues
Authors: Fisher John E. , Burger Peter C. , Perlman Elizabeth J. , Dickman Paul S. , Parham David M. , Savell Van H. , Hutchison Robert E. , Paidas Charles N. , Lev Elaine Rappaport
Published: July 2002

This article is the offshoot of a Pediatric Oncology Group (POG) seminar presented at the Adams Mark Hotel, Denver, Colorado, Friday, May 21, 1999, titled “The Frozen Section in Pediatric Solid Tumors—Crucial Issues.” There were eight presenters who spoke on a wide range of topics that included historical perspectives of the frozen section and discussion of the following systems: brain, renal, germ cell, bone, soft tissue, and lymph nodes. To complement these presentations, a pediatric surgeon explained his concern and philosophy regarding the use of frozen sections, and a lawyer tackled the issues and risks in rendering a frozen section diagnosis. We think that this review covers all the important aspects of the frozen section in our current practice of pediatric pathology.
Identification of a calmodulin‐dependent glycogen synthase kinase in rabbit skeletal muscle, distinct from phosphorylase kinase
Authors: Woodgett James R. , Tonks Nicholas K. , Cohen Philip
Published: July 2002

A glycogen synthase kinase that is completely dependent on Ca2+ and calmodulin has been identified in mammalian skeletal muscle, and purified ∼3000‐fold by chromatography on phosphocellulose and calmodulin—Sepharose. The presence of 50 mM NaCl in the homogenisation buffer was critical for extraction of the enzyme. The calmodulin‐dependent glycogen synthase kinase (app. M
r 850 000) is distinct from myosin light‐chain kinase and phosphorylase kinase, but phosphorylates the same serine residue on glycogen synthase as phosphorylase kinase. The physiological role of the enzyme is discussed.
Formation of the COOH‐terminal amide group of thyrotropin‐releasing‐factor
Authors: Husain Intisar , Tate Suresh S.
Published: July 2002

The precursors of peptide hormones that possess a COOH‐terminal α‐amide group contain a glycine residue following the amino acid which is amidated in the hormone. 〈Glu—His—Pro—Gly was synthesized as a putative precursor of thyroliberin. Bovine pituitary neurosecretory granules were shown to contain an amide group‐forming activity associated with an M
r of about 62000 protein(s) which converts the tetrapeptide to thyroliberin.
Specific DNA binding of the cyclic AMP receptor protein to a synthetic oligodeoxyribonucleotide
Authors: Martin Stephen R. , Gronenborn Angela M. , Clore G.Marius
Published: July 2002

The interaction of the cAMP receptor protein (CRP) of Escherichia coli with a synthetic DNA undecamer (11mer) comprising a portion of the specific target site in the gal operon and containing 8 basepairs out of the 10 basepair concensus making up specific CRP sites, has been studied by circular dichroism spectroscopy. The binding constants for the interaction of CRP with the 11mer in the presence and absence of cAMP have been determined, and it is shown that CRP, both in the presence and absence of cAMP, induces a B—C transition in the conformation of the 11mer.
An examination of the basic blue copper protein from cucumber peelings by 1H‐NMR
Authors: King G. , Andary T.A. , Freeman H.C. , Gavrilovic L. , Wright P.E.
Published: July 2002

The 1H‐NMR spectrum of cucumber basic blue protein (CBP) has been recorded. Examination of the spectrum of the reduced protein suggests that one or more sidechains exist in conformations which interconvert slowly at ambient temperatures. His 39, His 84 and Met 89 are identified as copper ligands by redox titration and by amino acid sequence homology with plastocyanin and azurin. The importance of a Phe sidechain close to the Met ligand in the potential blue copper site is confirmed. Broadening of His ligand resonances at elevated temperatures reveals an exchange process at the reduced copper centre.
Deoxythymidine pools of human skin and guinea pig organs
Authors: Harmenberg J. , Malm M. , Abele G.
Published: July 2002

The activity of antiviral nucleoside analogues like acyclovir is influenced by a number of cellular factors, one being the deoxythymidine (dThd) concentration. We therefore analysed the dThd concentration in human plasma and skin and in organs of guinea pig, the common experimental animal. High‐performance liquid chromatography showed low amounts of dThd in human skin, 0.20–1.15 , whereas guinea pig skin and spleen had 20–30 and the concentration in guinea pig plasma was 10‐times higher than in human plasma. These animals are therefore in this respect less suitable as accurate models for antiviral nucleoside activity in humans.
Characterization of the chymotryptic core of the adenovirus DNA‐binding protein
Authors: Tsernoglou D. , Tsugita A. , Tucker A.D. , van der Vliet P.C.
Published: July 2002

A fragment of the DNA‐binding protein of adenovirus type 5 has been obtained by controlled chymotryptic digestion of the entire molecule. Partial sequence determination indicates that the fragment consists of amino acids 174–525. The fragment is biologically active as measured by its ability to substitute for the entire molecule in a reconstituted DNA replication system. Crystals have been obtained that show diffraction to 2 Å.
Sirohydrochlorin, a precursor of factor F430 biosynthesis in Methanobacterium thermoautotrophicum
Authors: Mucha Helmut , Keller Eberhard , Weber Hans , Lingens Franz , Trösch Walter
Published: July 2002

Factor F430 is a nickel‐containing coenzyme of methanogenic bacteria with porphinoid structure which is derived from uroporphyrinogen III. It is shown that sirohydrochlorin is metabolized by cell free extracts of Methanobacterium thermoautotrophicum to factor F430 demonstrating that this compound, or a reduced form of it, is an intermediate in the biosynthesis of F430, and not only of vitamin B12 and siroheme.
Birhythmicity in a model for the cyclic AMP signalling system of the slime mold Dictyostelium discoideum
Authors: Goldbeter Albert , Martiel Jean-Louis
Published: July 2002

We demonstrate the coexistence of two simultaneously stable periodic regimes in a model based on receptor desensitization for the cyclic AMP signalling system of the slime mold Dictyostelium discoideum. These results provide the first example of autonomous birhythmicity in a biochemical model closely related to experimental observations. Whereas the transition from one stable mode of oscillations to the other can be elicited by suprathreshold stimuli, the two periodic regimes differ in their sensitivity to perturbations. That multiple oscillations occur in a model based on a single feedback loop suggests that the conditions for birhythmicity are widely satisfied in biological systems.
The structure of C hordein
Authors: Field J.M. , Tatham A.S. , Baker A.M. , Shewry P.R.
Published: July 2002

The intrinsic viscosity of C hordein was determined in 70% () aqueous ethanol (± 10 mM NaCl) and 0.1 M acetic acid at temperatures between 15.3 and 50°C. Calculations based on the assumption (supported by previously published results) that C hordein has an ordered conformation indicated that the molecule is rod shaped, with molecular dimensions varying from about 360 × 17 Å to 265 × 20 Å. Circular dichroism spectroscopy indicated a secondary structure rich in β‐turns, with some conformational differences in the two solvents and at different temperatures. The results are compatible with C hordein having a helical secondary structure based on repetitive β‐turns.
Cloning of rat brain protein kinase C complementary DNA
Authors: Ono Yoshitaka , Kurokawa Tsutomu , Kawahara Kenji , Nishimura Osamu , Marumoto Ryuji , Igarashi Koichi , Sugino Yukio , Kikkawa Ushio , Ogita Kouji , Nishizuka Yasutomi
Published: July 2002

Four peptides derived from rat brain protein kinase C were partially sequenced. Using synthetic oligonucleotides deduced from the amino acid sequences as probes, a clone of complementary DNA (cDNA) was isolated from a cDNA library prepared from the same tissue. The nucleotide sequence of this cDNA clone revealed the primary structure of the carboxyl‐terminal region as having 224 amino acids, with significant sequence homology with cyclic AMP‐dependent and cyclic GMP‐dependent protein kinases.
Characterisation of phosphate binding to mitochondrial and bacterial membrane‐bound ATP synthase by studies of inhibition with 4‐chloro‐7‐nitrobenzofurazan
Authors: Perez Juan A. , Greenfield Anthony J. , Sutton Raul , Ferguson Stuart J.
Published: July 2002

The effect of phosphate on the inhibition by 4‐chloro‐7‐nitrobenzofurazan of the ATPase activity of the proton‐translocating ATP synthase in heart submitochondrial particles was investigated. Binding of phosphate protected strongly against the inhibition. A dissociation constant of 0.2 mM was determined for the enzyme · Pi complex and shown to be independent of pH in the range 7.0–8.0. The protective effect of phosphate was mimicked by arsenate but not by sulphate or malonate. Similar results were obtained for the enzyme from Paracoccus
denitrificans. 2,4‐Dinitrophenol enhanced phosphate binding to the mitochondrial enzyme since the protective effect of phosphate was increased. The data are compatible with protection arising from binding of phosphate to a catalytic site.
A model of the nucleotide‐binding site in tubulin
Authors: Sternlicht Himan , Yaffe Michael B. , Farr George W.
Published: July 2002

Tubulin uses GTP to regulate microtubule assembly and is thought to be a member of a class of GDP/GTP‐binding proteins (G‐proteins) as defined by Hughes [(1983) Febs Lett. 164, 1–8]. How tubulin is structurally related to G‐proteins is not known. We use a synthesis of sequence comparisons between tubulin, other G‐proteins, and ADP/ATP‐binding proteins and topological arguments to identify potential regions involved in nucleotide binding. We propose that the nucleotide‐binding domain in the β‐subunit of tubulin is an α/β structure derived from amino acid residues ∼60–300. Five peptide sequences are identified which we suggest exist as ‘loops’ that extend from β‐strands and connect α‐helices in this structure. We argue that GDP binds to four of the five loops in an Mg2+‐independent manner while GTP binds in an Mg2+‐dependent manner to a different combination of four loops. We propose that this switch between loops upon GTP binding induces a conformational change essential for microtubule assembly.
Cleavage of the ArgIle bond in the native polypeptide chain of human pancreatic stone protein
Authors: Rouimi P. , Bonicel J. , Rovery M. , De Caro A.
Published: July 2002

The pancreatic stone protein (PSP) isolated from calculi (M
r 14000) and the 5 protein forms (PSP S1‐5) detected in pancreatic juice (M
r 14000—19000) derive from the same source differing seemingly in their carbohydrate contents or/and in their polypeptidic chain lengths. This kind of protein would inhibit in vivo CaCO3‐crystal growth in pancreatic juice. PSP and PSP S1 N‐terminal sequences are identical (NH2Ile‐). This report demonstrates that: (i) in PSP S2‐5 the amino‐terminal is blocked; (ii) the C‐terminus is alike in every form; (iii) the single polypeptide chain of PSP S2‐5 is converted into that of PSP S1 or PSP by the specific trypsin cleavage of the ArgIle bond.
Femtosecond spectroscopy of primary charge separation in modified reaction centers of Rhodobacter sphaeroides (R‐26)
Authors: Chekalin S.V. , Matveetz Ya.A. , Shkuropatov A.Ya. , Shuvalov V.A. , Yartzev A.P.
Published: July 2002

Femtosecond measurements of kinetics and spectra of absorbance changes (ΔA) were carried out with modified reaction centers (RCs) from Rhodobacter sphaeroides (R‐26) from which nonactive bacteriochlorophyll BM (located in the M protein subunit) was removed. The band of BM at 800 nm in native RCs is shifted in femtosecond measurements and obscures the ΔA of active bacteriochlorophyll BL (L subunit). The spectrum of ΔA in modified RCs at 6 ps delay includes the bleachings of the bands of P (primary electron donor) at 870 nm, of BL at 805 nm and of HL (bacteriopheophytin located in the L subunit) at 755 nm showing the reduction of 0̃.5 mol BL and 0̃.5 mol HL per mol P+. These data confirm an earlier suggestion that BL participates as an electron acceptor in the light‐induced primary charge separation and agree with recent X‐ray analysis of Rhodopseudomonas viridis and R. sphaeroides RCs which shows a location of BL between P and HL.
Enkephalin‐degrading enzymes and angiotensin‐converting enzyme in human and rat meninges
Authors: Zajac Jean-Marie , Charnay Yves , Soleilhac Jean-Marc , Sales Nicole , Roques Benard P.
Published: July 2002

The neutral endopeptidase NEP 24.11 (enkephalinase) has been visualized in human spinal cord by in vitro autoradiography using [3H]HACBO‐Gly as a radiolabelled probe. The specific binding was present in the substantia gelatinosa and particularly dense in meninges surrounding the spinal cord. Enzymatic studies using [3H][D‐Ala2, Leu]enkephalin as substrate confirmed the presence of NEP in dura and pia mater of human tissue. In addition, the human meninges were shown to contain high concentrations of angiotensin‐converting enzyme (ACE) and aminopeptidases. The three enzymes have also been detected in rat tissues but their distribution pattern differs from that of human tissue. In dura mater, 45% of the [Leu]enkephalin hydrolysis was due to enkephalinase and 38% to bestatin‐sensitive aminopeptidases. In contrast in pia mater aminopeptidases were more efficient in hydrolyzing enkephalin. The possible role of these enzymes in the meninges could be to maintain the homeostatic concentration of neuropeptides in the central nervous system.
Ca2+/calmodulin‐dependent protein kinase in Saccharomyces cerevisiae
Authors: Londesborough John , Nuutinen Marja
Published: July 2002

Ca2+‐dependent chromatography of soluble cytosolic proteins on calmodulin‐Sepharose gave a fraction that exhibited Ca2+‐ and calmodulin‐dependent phosphorylation of several polypeptides, including 60, 56 and 45 kDa species. At 0.2 μM beef calmodulin the phosphorylation was optimal at 3 μM free Ca2+, and at 80 μM free Ca2+ it was half‐maximal at about 0.1 μM beef calmodulin. It is concluded that the fraction contains calmodulin‐dependent protein kinase(s) which is (are) autophosphorylated or associated with substrates.
Changes in the proportions of histone H1° subtypes in brain cortical neurons
Authors: Piña B. , Suau P.
Published: July 2002

Histone H1° is found in tissues with little or no cellular proliferation and has been shown to accumulate during cellular terminal differentiation. Two subtypes of H1°, H1°a and H1°b, are present in any tissue where the protein has been detected. We report here the first evidence of an age‐dependent change in the proportions of H1° subtypes. In rat cerebral cortex neurons the proportion of H1°a rises from 44% of total H1° at birth to about 80% at day 300. These results show that terminally differentiated neurons synthesize and exchange H1° at a significant rate.
Discrete extraction of the Ca atom functional for O2 evolution in higher plant photosystem II by a simple low pH treatment
Authors: Ono Taka-aki , Inoue Yorinao
Published: July 2002

Treatment of spinach PS II membranes with a citrate solution at pH 3.0 totally inactivated O2 evolution concomitant with a 50% decrease in Ca abundance. Notably, neither the abundance of Mn and extrinsic proteins nor the activity of DPC photooxidation was at all affected by the treatment. The treated membranes evolved O2 at a high rate in the presence of exogenous Ca2+, but the activity was sensitive to EDTA. However, when the treated membranes were incubated with Ca2+ for a few tens of minutes, the O2‐evolving activity became EDTA‐resistant, suggesting a firm re‐ligation of Ca2+ to the Ca‐binding site. It was indicated that spinach PS II contains two Ca atoms per reaction center, and that the low pH citrate treatment selectively removes one of the two Ca atoms that is specifically functional for O2 evolution, even in the presence of all three extrinsic proteins.
Conformational changes of type II regulatory subunit of cAMP‐ dependent protein kinase on cAMP binding
Authors: Johnson Dennis R. , Wong Shan S.
Published: July 2002

The effect of cAMP on the conformation of the regulatory subunit of type II cAMP‐dependent protein kinase (RII) from bovine heart was investigated by UV‐difference and circular dichroism (CD) spectroscopy. The UV‐difference spectrum of RII with and without cAMP showed a positive band around 278 nm and a negative band at 256 nm. Similarly, cAMP enhanced the ellipticity of RII in the region between 280 and 300 nm and decreased that between 250 and 280 nm. In addition, cAMP transformed the far‐UV CD spectrum of RII from that of a negative band minimally at 209 nm with a shoulder at 223 nm to one with two minima at 222 and 211 nm. These data show that cAMP induces conformational changes of RII upon binding. Such structural changes may be the basis of activation of cAMP‐dependent protein kinases by cAMP.
Processing of Ada protein by two serine endoproteases Do and So from Escherichia coli
Authors: Lee Cheol Soon , Hahm Jae Kyu , Hwang Byung Joon , Park Kyung Chan , Ha Doo Bong , Park Sang Dai , Chung Chin Ha
Published: July 2002

Two soluble serine proteases Do and So from Escherichia coli were found to distinctively cleave the purified, 39 kDa Ada protein into fragments with sizes of 12–31 kDa. Protease So appears to generate a C‐terminal 19 kDa polypeptide, similarly to OmpT protease. In addition, the purified 19 kDa C‐terminal half of Ada protein can be further processed mainly to an 18 kDa fragment by protease So and to a 12 kDa by protease Do. These results suggest that proteases Do and So are involved in endogenous cleavage of Ada protein, which may play a role in down‐regulating the adaptive response to alkylating agents.
Oxidation of non‐phenolic substrates
Authors: Bourbonnais Robert , Paice Michael G.
Published: July 2002

In the presence of substrates such as Remazol Blue and 2,2'‐azinobis(3‐ethylbenzthiazoline‐6‐sulphonate) (ABTS), laccases Coriolus (Trametes)
versicolor can also oxidize non‐phenolic lignin model compounds. Veratryl alcohol (I) and 1‐(3,4‐dimethoxyphenyl)‐2‐(2‐methoxyphenoxy)‐propane‐1, 3‐diol (III) were oxidized by laccase and mediator to give the α‐carbonyl derivatives. The β‐1 lignin model dimer, 1‐(3,4‐dimethoxyphenyl)‐2‐phenoxy‐ethane‐1, 2‐diol(II) was cleaved by laccase in the presence of ABTS to give veratraldehyde and benzaldehyde. On the basis of these observations, we propose that laccase is capable of oxidizing both phenolic and non‐phenolic moieties of lignin but that the latter is dependent on the co‐presence of primary laccase substrates.
N‐Furfurylformamide as a pseudo‐substrate for formylmethanofuran converting enzymes from methanogenic bacteria
Authors: Breitung J. , Börner G. , Karrasch M. , Berkessel A. , Thauer R.K.
Published: July 2002

Methanofuran involved in methanogenesis. The N‐formyl derivative is an intermediate in the reduction of CO2 to CH4 and the disproportionation of methanol to CO2 and CH4. Formylmethanofuran dehydrogenase and formylmethanofuran:tetrahydromethanopterin formyltransferase are the enzymes catalyzing its conversions. We report here that the two enzymes from Methanosarcma barkeri and the formyltransferase from Methanobacterium thermoautotrophicum can also use N‐furfurylfonnamide as a pseudo‐substrate albeit with higher apparent K
m and lower apparent V
max values. N‐Methylformamide, formamide, and formate were not converted indicating that the furfurylamine moiety of methanofuran is the minimum structure required for the correct binding of the coenzyme.
Inhibition of poly(ADP‐ribose) synthetase by unsaturated fatty acids, vitamins and vitamin‐like substances
Authors: Banasik Marek , Komura Hajime , Ueda Kunihiro
Published: July 2002

Various vitamins and vitamin‐like substances inhibited the activity of poly(ADP‐ribose) synthetase in vitro. The most potent were essential fatty acids, i.e. arachidonic acid, linoleic acid, and linolenic acid; their 50% inhibitory concentrations (IC50) were 44–110 μM, indicating a higher potency than nicotinamide, a well‐known vitamin inhibitor (IC50 = 210 μM). Vitamins K3, K1, and retinal were the next strongest inhibitors, followed by α‐lipoic acid, coenzyme Q10, and pyridoxal 5‐phosphate. Nicotinamide and vitamin K3, exhibited mixed‐type inhibition with respect to NAD+,while arachidonic acid exhibited dual inhibitions, competitive at 50 μM and mixed‐type at 100 μM.
Mechanisms of differential regulation of interleukin‐6 mRNA accumulation by tumor necrosis factor alpha and lymphotoxin during monocytic differentiation
Authors: Brach Marion A. , Cicco Nicola A , Riedel Detlev , Hirano Toshio , Kishimoto Tadamitsu , Mertelsmann Roland H. , Herrmann Friedhelm
Published: July 2002

In the present report we compare the capacity of two related cytokines, tumor necrosis factor (TNF) alpha and lymphotoxin (LT), to modulate mRNA levels of interleukin‐6 (IL‐6) in cells representing different stages of monocytic differentiation including the human leukemia cell lines HL 60, U 937, THP‐1, MonoMac 1 and peripheral blood monocytes. We show that the capacity of TNF alpha and LT to induce IL‐6 mRNA accumulation increases as monocytic differentiation proceeds with TNF alpha being more potent than LT, suggesting that alternate pathways may be used by differentiating cells to control expression of IL‐6. In contrast, in monocytes which constitutively synthesize IL‐6 transcripts, TNF alpha and LT treatment had opposite effects on levels of IL‐6 mRNA accumulation. In these cells TNF alpha enhanced steady state levels of IL‐6 transcripts due to mRNA stabilization, whereas LT shortened IL‐6 mRNA half‐life, most likely due to induction of a RNA destabilizer since LT‐mediated downregulation of levels of IL‐6 mRNA in monocytes could be prevented by inhibition of protein synthesis. Neither TNF alpha nor LT altered IL‐6 mRNA accumulation by interfering with preexisting transcription factors since both TNF alpha and LT required de novo protein synthesis to exert their effects.
Tryptase TL2 in the membrane of human T4+ lymphocytes is a novel binding protein of the V3 domain of HIV‐1 envelope glycoprotein gp120
Authors: Kido Hiroshi , Fukutomi Aiko , Katunuma Nobuhiko
Published: July 2002

A novel membrane‐bound serine esterase in cultured human T4+ lymphocytes, recently purified and named tryptase TL2, binds specifically to the external envelope protein gp12O of HIV‐1, interacting with its V3 domain. This binding was selectively blocked by inhibitors of tryptase TL2 with a GPCR sequence in their reactive site, synthetic peptides corresponding with the sequences of the V3 domains of various HIV‐1 strains with the GPGR sequence, and antibody against tryptase TL2, or neutralizing antibody against the V3 domain of HTLV‐IIIB. These findings suggest that tryptase TL2 is a binding protein of the V3 domain of HIV‐1 envelope glycoprotein.
The phospholipid analogue hexadecylphosphocholine inhibits phosphatidylcholine biosynthesis in Madin—Darby canine kidney cells
Authors: Haase Rüdiger , Wieder Thomas , Geilen Christoph C. , Reutter Werner
Published: July 2002

The influence of the phospholipid analogue hexadecylphosphocholine on phosphatidylcholine biosynthesis was investigated in Madin—Darby canine kidney (MDCK) cells. It inhibits the incorporation rate of [methyl‐3H]choline into phosphatidylcholine at a concentration of 50 μM by about 50%. The radiolabelled precursor accumulates in the phosphocholine pool indicating that hexadecylphosphocholine inhibits the formation of phosphatidylcholine via the CDP‐choline pathway at the level of the rate‐limiting enzyme, CTP:phosphocholine cytidylyl transferase (EC 2.7.7.15). This was verified by the determination of the activity of the enzyme in vitro. In consequence of its inhibitory effect it could be shown that the treatment of MDGK cells for 24 h with 50 μM hexadecylphosphocholine induces alterations of the phospholipid composition. Whereas in treated cells the relative phosphatidylcholine content was decreased from the control level of 36.0±0.9% to 29.9±0.2%; in contrast, the relative content of phosphatidylethanolamine was increased from 19.3±0.9% to 24.3±0.9%.
Apamin‐sensitive potassium channels mediate agonist‐induced oscillations of membrane potential in pituitary gonadotrophs
Authors: Kukuljan Manuel , Stojilković Stanko S. , Rojas Eduardo , Catt Kevin J.
Published: July 2002

In cultured rat pituitary gonadotrophs, gonadotropin‐releasing hormone (GnRH) induces rapid hyperpolarization of the cell membrane and causes cessation of the spontaneous electrical activity present in non‐stimulated cells. This initial response to GnRH is followed by slow oscillations of membrane potential (V
m) which often exhibit brief bursts of action potentials (AP) fired from the peak of the oscillations. The hyperpolarization waves are synchronous with GnRH‐induced elevations of cytoplasmic Ca2+ concentration ([Ca2+]i), such that V
m maxima alternate with the peak values of [Ca2+]i. The V
m oscillations result from repetitive activation of apamin‐sensitive K+ channels by cytoplasmic Ca2+. Thus, GnRH activation of Ca2+ mobilization can generate a bursting pattern of membrane potential through the activation of K+ channels against a background of spontaneous electrical activity.
Putative nucleotide binding sites of guinea pig liver transglutaminase
Authors: Takeuchi Y. , Birckbichler P.J. , Patterson M.K. , Lee K.N.
Published: July 2002

Three peptides corresponding to glycine‐rich internal sequences of the guinea pig liver transglutaminase molecule were synthesized. These were peptide 1 (amino acid residues 520–544), peptide 2 (amino acid residues 345–367) and peptide 3 (amino acid residues 45–69). All of the synthetic peptide demonstrated significant binding ability for both ATP and GTP. Peptide 1 was the best protector or transglutaminase activity from both ATP and GTP inhibition, while peptides 2 and 3 protected the activity only from GTP inhibition. The data shown here lead us to propose putative binding site(s) for ATP and GTP guinea pig liver transglutaminase.
Specific release of a 9‐kDa extrinsic polypeptide of photosystem I from spinach chloroplasts by salt washing
Authors: He Wei-Zhong , Malkin Richard
Published: July 2002

The newly reported 9‐kDa polypeptide in photosystem I [(1991) FEBS Lett. 280, 332–334) is an extrinsic component located on the lumenal side of the thylakoid membrane. This subunit can be solubilized with high salt buffer and does not bind any cofactors. The photosystem I electron transfer chain remains intact and functional in the absence of this component as characterized by the photoreduction of NADP*.
The inverted hexagonal phase is more sensitive to hydroperoxidation than the multilamellar phase in phosphatidylcholine and phosphatidylethanolamine aqueous dispersions
Authors: Ye Wang Jin , Miyazawa Teruo , Fujimoto Kenshiro , Wang ZhengYu , Nozawa Tsunenori
Published: July 2002

The effect of phase behaviour (hexagonal II phase and lamellar phase) on the peroxidation of membrane phospholipids has been investigated in dilinoleoyl phosphatidylcholine (DLPC)/dilinoleoyl phosphatidylethanolamine (DLPE) aqueous dispersions. Peroxidation was initiated with a water‐soluble radical inducer 2,2′‐azobis (2‐amidino‐propane) dihydrochloride (AAPN). The phospholipid morphology was monitored by 31P‐nuclear magnetic resonance (NMR). Phospholipid hydroperoxides (PCOOH and PEOOH) were determined by chemiluminescence high‐performance liquid chromatography (CL—HPLC). In pH‐induced phase transition systems, DLPE in the bilayer state was much less oxidized than in the hexagonal II state. In composition‐induced phase transition systems, the formation of total hydroperoxides and the consumption of α‐tocopherol in the hexagonal II phase were greater than in the bilayer phase. These data suggest that the hexagonal II phase is more sensitive to hydroperoxidation than the bilayer phase in phospholipid aqueous dispersions.
On the production of α,β‐heterodimeric acyl‐coenzyme A: isopenicillin N‐acyltransferase of Penicillium chrysogenum
Authors: Aplin Robin T. , Baldwin Jack E. , Cole Stephen C.J. , Sutherland John D. , Tobin Matthew B.
Published: July 2002

A high level E. coli expression system has been constructed for the Penicillium chrysogenum penDE gene, which encodes the acyl‐coenzyme A: isopenicillin N‐acyltransferase (AT) enzyme. Induction of overexpression of recombinant AT (recAT) by increasing the growth temperature of the host adversely affected solubility and activity of the AT enzyme. Addition of isopropylthio‐β‐d‐galactopyranoside (IPTG) at decreased growth temperatures (less than 32°C) resulted in the overproduction of soluble, active recAT. When purified to homogeneity, recAT was an α,β‐heterodimer, comprised of 11 kDa (α) and 29 kDa (β) subunits, derived from a 40 kDa precursor polypeptide by a posttranslational cleavage. The recAT enzyme contained both the acyl‐coenzyme A: isopenicillin N‐acyltransferase and the acyl‐coenzyme A: 6‐aminopenicillanic acid acyltransferase activities. The processing event that generated the two subunits of recAT from the 40 kDa precursor polypeptide occurred between Gly102/Cys103. This expression system produced a large amount of soluble, active recAT that is identical to native AT, making it a suitable source of AT enzyme for further characterization.
Tyrphostin‐induced differentiation of mouse erythroleukemia cells
Authors: Anafi Mordechai , Gazit Aviv , Gilon Chaim , Neriah Yinon Ben , Levitzki Alexander
Published: July 2002

Inhibitors of protein‐tyrosine kinases (TPKs) from the tyrphostins family induce terminal erythroid differentiation of mouse erythroleukemia (MEL) cells. The most potent tyrphostin was found to be AG‐568 which was therefore investigated in more detail. Just prior to differentiation the inhibition of tyrosine phosphorylation of a pp97 protein band was noted. We also found that AG‐568 treatment induces the appearance of a putative differentiation factor which could induce tyrphostin‐independent differentiation in untreated cells. Our study suggests that the inhibition of tyrosine phosphorylation by AG‐568 leads to the production of differentiating factor(s) which induce the MEL cells to differentiate.
Stoichiometry of the binding of chromosomal protein MCl from the archaebacterium, Methanosarcina spp. CHTI55, to DNA
Authors: Culard Françoise , Laine Bernard , Sautiére Pierre , Maurizot JeanClaude
Published: July 2002

We have investigated the binding Stoichiometry of the chromosomal MCl protein on DNA using the gel retardation technique. Analysis of the distribution of the complex containing 0, 1, 2, 3...... bound proteins shows that the protein MCl interacts with the DNA as a monomer. Binding experiments with short DNA fragments of various lengths shows that the site size is 11 bp in length. These results are compared to those obtained with other chromosomal proteins including HU protein.
A negative charge in the M2 transmembrane segment of the neuronal α7 acetylcholine receptor increases permeability to divalent cations
Authors: Ferrer-Montiel Antonio V. , Montal Mauricio
Published: July 2002

Threonine‐244 (T244) in the putative channel‐forming M2 segment of the neuronal α7 acetylcholine receptor (AChR), a residue proposed to form part of the selectivity filter, was mutated to aspartic acid to examine the influence of a negative charge on AChR ion permeation properties. Wild type (AChRα7wt) and mutant (AChRα7D244) acetylcholine receptors expressed in Xenopus oocytes give rise to acetylcholine (ACh)‐activated, α‐bungarotoxin‐sensitive, cation‐selective ionic currents. AChRα7D244 exhibited larger currents than AChRα7wt that, in addition, activated at lower ACh concentrations. The relative ionic permeability (P
x/P
Na) of AChRα7wt to K+ was P
K/P
Na = 1.2, and to Ba2+, P
Ba/P
Na = 1.4. In contrast, AChR α7D244 was less selective in discriminating between K+ and Na+, P
k/P
Na = 0.95, but exhibited a remarkable increase in permeability to Ba2+, P
Ba/P
Na = 3.7. Furthermore, only mutant receptors were permeable to Mg2+. Hence, a ring of negatively charged residues in the putative pore‐forming segment of the receptor increases the permeability to divalent cations. Our results substantiate the notion that T244, or its equivalent, in the M2 transmembrane segment of cholinergic receptor channels is a key structural determinant of the selectivity filter.
Two 3',5'‐cyclic‐adenosine monophosphate response elements in the promoter region of the human gastric inhibitory polypeptide gene
Authors: Someya Yoshimichi , Inagaki Nobuya , Maekawa Toshio , Seino Yutaka , Ishii Shunsuke
Published: July 2002

Transfection of chimeric chloramphenicol acetyltransferase plasmids containing various deletions of the human gastric inhibitory polypeptide (GIP) promoter into hamster insulinoma (HIT T15) cells indicated that the region between −180 and +14 is sufficient for basal promoter activity. Two CRE‐BP1 binding sites were identified in this promoter region by DNase I footprinting with the bacterially expressed cAMP response element (CRE) binding protein, CRE‐BP1. Mutation analyses showed that these two CREs are required for the basal promoter activity, and furthermore that one site, at nucleotide‐158, contributed mainly to the cAMP inducibility of the GIP promoter in HIT T15 cells. Interestingly, the GIP promoter activity was repressed by the c‐jun proto‐oncogene product, possibly through the CREs.
Three‐dimensional structural resemblance between the ribonuclease H and connection domains of HIV reverse transcriptase and the ATPase fold revealed using graph theoretical techniques
Authors: Artymiuk Peter J. , Grindley Helen M. , Kumar Kiran , Rice David W. , Willett Peter
Published: July 2002

Using 3D searching techniques based on algorithms derived from graph theory, we have established two previously unreported structural similarities involving the ribonuclease H (RNase H) domain of HIV‐1 reverse transcriptase (RT). First, we report that there is a strong similarity between the 3D folds of the RNase H domain of RT and the ‘ATPase folds’ of hexokinase, the 70 kDa heat‐shock cognate protein and actin. Like RNase H, these enzymes are involved in nucleotide binding and metal ion‐catalysed cleavage of a phosphodiester bond. Similarities of the folding motif and the position of the metal‐binding site in these enzymes suggest possible functional analogies and evolutionary relationships with RNase H. Second, we find there is a strong resemblance between the folds of the RNase H domain and of the p66 and p51 ‘connection’ domains of RT. It is possible that this striking similarity within the RT structure indicates a possible ancestral gene doubling event. The similarity may also indicate that the connection domains possess functional roles in addition to those previously suggested, and they may therefore represent a further target for the design of therapeutic agents.
A parallel three stranded α‐helical bundle at the nucleation site of collagen triple‐helix formation
Authors: Hoppe Hans-Jürgen , Barlow Paul N. , Reid Kenneth B.M.
Published: July 2002

A short stretch of 35 amino acids is identified as the structural motif responsible for the tight parallel association and trimerization of the three identical polypeptide chains of lung surfactant protein D, which contains both collagen regions and C‐type lectin domains. This ‘neck‐region’ is located at the nucleation site at which the collagenous sequences fold into a staggered triple‐helix and is shown, by CD, NMR, and cross‐linking of recombinant peptides, to consist of a triple‐stranded parallel α‐helical bundle in a non‐staggered, and extremely strong, non‐covalent association. This type of association between three polypeptide chains may represent a common structural feature immediately following the C‐terminal end of the triple‐helical region of collagenous proteins.
Sulfatides trigger cytokine gene expression and secretion in human monocytes
Authors: Constantin Gabriela , Laudanna Carlo , Baron Pierluigi , Berton Giorgio
Published: July 2002

We investigated whether sulfatides are able to trigger transmembrane signals and activation of selective cell functions in human monocytes. Sulfatides stimulated an increase in cytosolic free‐calcium in monocytes, and this depended on the release of calcium from intracellular stores. Non‐sulfated galactocerebrosides had no effect on monocyte cytosolic free calcium. Sulfatides enhanced expression of tumor necrosis factor, interleukin‐8, and interleukin‐1β, but not interleukin‐12/natural killer cell stimulating factor mRNAs. Sulfatides also triggered secretion of cytokines into the extracellular medium, although they were much less effective than lypopolysaccharide. Both enhanced expression of cytokine mRNAs and secretion by sulfatides required sulfation of the galactose ring of the glycolipid as non‐sulfated galactocerebrosides had no effect. These findings suggest that sulfatides that are released at sites of inflammation can amplify the inflammatory reaction triggering cytokine expression in, and release by, monocytes.
A MGF/STAT5 binding site is necessary in the distal enhancer for high prolactin induction of transfected rabbit αs1‐casein‐CAT gene transcription
Authors: Jolivet Geneviève , L'Hotte Corinne , Pierre Sandra , Tourkine Nikolaï , Houdebine Louis-Marie
Published: July 2002

The rabbit αs1‐casein gene contains a distal prolactin‐dependent enhancer 3442‐3285 bp 5′ to the site of initiation of transcription. We have reported previously that four DNA/protein‐binding sites (F1–F4) are located within this distal enhancer. We now show that one of this binding site (the F4 site) binds in vitro a MGF/STAT5‐like factor. The functional importance of the F4 site was estimated by cotransfection of CHO cells with a chimeric gene containing or not the F4 sequence linked to the (−391/+1774)CAT gene and a plasmid encoding the rabbit mammary prolactin receptor. The F4 site is necessary for maximal response of the enhancer to prolactin. However, this site has to be associated to the F1–F3 fragment. It can be replaced by a genuine MGF/STAT5‐binding site. A mutational analysis indicates that F4 and F1 sites are simultaneously involved to confer a high prolactin sensitivity.
NADPH:protochlorophyllide oxidoreductase from Synechocystis: overexpression, purification and preliminary characterisation
Authors: Heyes Derren J. , Martin Giles E.M. , Reid R.James , Hunter C.Neil , Wilks Helen M.
Published: July 2002

NADPH:protochlorophyllide oxidoreductase (POR) catalyses the light‐dependent reduction of protochlorophyllide to chlorophyllide, a key regulatory reaction in the chlorophyll biosynthetic pathway. POR from the cyanobacterium Synechocystis has been overproduced in Escherichia coli with a hexahistidine tag at the N‐terminus. This enzyme (His6‐POR) has been purified to homogeneity and a preliminary characterisation of its kinetic and substrate binding properties is presented. Chemical modification experiments have been used to demonstrate inhibition of POR activity by the thiol‐specific reagent N‐ethyl maleimide. Substrate protection experiments reveal that the modified Cys residues are involved in either substrate binding or catalysis.
Fenofibrate prevents and reduces body weight gain and adiposity in diet‐induced obese rats
Authors: Mancini F.P , Lanni A , Sabatino L , Moreno M , Giannino A , Contaldo F , Colantuoni V , Goglia F
Published: July 2002

Fibrates are hypolipidemic drugs that activate the peroxisome proliferator‐activated receptors. Since fibrates may also increase energy expenditure, we investigated whether fenofibrate (FF) had this effect in diet‐induced obese rats. A 2‐month administration of a high‐fat palatable diet to adult rats increased body weight by 25% and white adipose mass by 163% compared with a standard diet. These effects were prevented by FF, both when administered for the 2 months of high‐fat feeding and when given for only the second month. Consequently, FF‐treated rats had a final body weight and white adipose tissue mass similar to untreated animals on the standard diet. FF also increased resting metabolic rate, hepatic peroxisomal and mitochondrial palmitoyl‐dependent oxygen uptake and mRNA levels of acyl‐CoA oxidase and lipoprotein lipase. Finally, FF lowered mRNA levels of uncoupling protein‐2 and did not affect mitochondrial respiration in skeletal muscle. Therefore, FF seems to act as a weight‐stabilizer mainly through its effect on liver metabolism.