bluuebunny/crossref_metadata_2025 huggingface dataset:
Superactivity and phase‐sensitivity of potato acid phosphatase entrapped in reverse micelles
Authors: Lalitha J. , Mulimani VH
Published: July 2007

Potato acid phosphatase, AcPase (E.C. 3.1.3.2) was entrapped in reverse micelles of cationic surfactant cetyltrimethylammonium bromide (CTAB) in isooctane and chloroform (1:1). The activity was studied at different values of Wo = ([water] / [surfactant]). AcPase exhibited superactivity in the reverse micellar system. At very low Wo value, activity was found to be less than that of in buffer and further increase in Wo value enhanced the activity thousand fold. At Wo=50, the activity was enhanced more than twenty fold. The effect of second surfactant, TritonX‐100 on superactivity was studeid. There was a slight decrease in overall activity, when 3.33 mM TritonX‐100 was added to the above reverse micellar system.
Elevated catalase activity in red and white muscles of MyoD gene‐inactivated mice
Authors: Tiidus Peter M. , Bombardier Eric , Xeni Jason , Bestic Nancy M. , Vandenboom Rene , Rudnicki Michael A. , Houston Michael E.
Published: July 2007

MyoD is a myogenic transcription factor responsible for skeletal muscle differentiation during development. Muscle antioxidant enzyme status was determined in transgenic MyoD deactivated mice. While catalase activity was significantly (P<0.05) elevated in soleus and extensor digitorum longus muscles from MyoD deactivated mice, superoxide dismutase and glutathione peroxidase activities were not. While this may imply a greater propensity for inherent oxidative stress, soleus glutathione status was similar between MyoD deactivated mouse and control soleus muscles. Catalase activity is localized primarily in peroxisomes. Therefore elevated catalase activity may also indicate the presence of factors associated with peroxisome proliferation in muscles from MyoD gene‐inactivated mice.
Evidence for the shikimate‐3‐phosphate interacting site in the N‐terminal domain of 5‐enolpyruvyl shikimate‐3‐phosphate synthase of Bacillus subtilis
Authors: Selvapandiyan Angamuthu , Ahmad Suhail , Majumder Kumud , Arora Naresh , Bhatnagar Raj K.
Published: July 2007

The role of basic amino acid residues that are highly conserved in the N‐terminal domain of 5‐enolpyruvyl shikimate‐3‐phosphate synthase (EPSPs) in the binding of the substrate, shikimate‐3‐phosphate, has been assessed. Lys 19 and Arg 24 in the Bacillus subtilis EPSPs were substituted by glutamic acid and aspartic acid residues respectively by site‐directed mutagenesis. Native and the mutant proteins were expressed using a two‐vector system and the expressed proteins were purified to near homogeniety. The replacement of either Lys 19 or Arg 24 with a negatively charged residue nearly completely abolished the enzyme activity. The kinetic characterization of the purified wild type and the mutant proteins revealed that the substitution of positively charged residues in the N‐terminal domain (K19 and R24) results in reduced affinity for shikimate‐3‐phosphate (S3P). The results suggest the involvement of these residues in the binding of S3P during enzyme catalysis.
GM‐CSF induces the tyrosine phosphorylation of three isoforms of shc and its association with GRB2 in TF‐1 cell
Authors: Shi Xiaoqing , Qin Junchuan , Zhu Dexu
Published: July 2007

This report demonstrates that GM‐CSF induces the tyrosine phosphorylation of Shc protein which is implicated in Ras activation. Three isoforms of She are ubiqitously phosphorylated induced by GM‐CSF in TF‐1, a cell line of erythroid origin. It is also shown that She is associated with the adaptor protein Grb2. The formation of Shc‐Grb2 complex may directly link tyrosine phosphorylation events to Ras activation in TF‐1 cells.
Bovine papilomavirus oncoprotein E5 induces the NFκB activation through superoxide radicals
Authors: Kilk Ann , Talpsepp Tiit , Väli Ülo , Ustav Mart
Published: July 2007

Bovine papillomavirus type 1 (BPV‐1) open reading frame E5 encodes the smallest known oncoprotein. This protein is involved in transforming cells during the early stage of infection. Nuclear factor κB (NFκB) is activated in the host cells as a result of different virus infections. It has been shown that reactive oxygen intermediates activate NFκB which leads to the activation of several cellular genes. We studied the effect of BPV‐1 E5 protein on NFκB activation. We found that low and moderate concentrations of E5 protein activate NFκB. This activation is suppressed by superoxide dismutase. We conclude that the superoxide anion is involved in activation of NFκB.
Ginsenoside‐Rh1 and Rh2 inhibit the induction of nitric oxide synthesis in murine peritoneal macrophages
Authors: Park Young Chul , Lee Chang Hoon , Kang Ho Sung , Kim Kyu‐Won , Chung Hun Taeg , Kim Han Do
Published: July 2007

The effects of ginsenoside‐Rh1 and Rh2 in the induction of nitric oxide (NO) synthesis by IFN‐γ plus LPS were investigated using murine peritoneal macrophages. The NO production from rIFNγ plus LPS‐treated macrophages was markedly reduced by ginsenoside‐Rh1 or Rh2 in a dose dependent manner, but had no inhibitory effects by ginsenoside‐Rb1, Rc or Re. In addition, treatment of the cells with ginsenoside‐Rh2 6 hr before the stimulation with IFN‐γ plus LPS showed more inhibitory effect than the treatment with ginsenoside‐Rh2 6 hr after or simultaneously with the stimulation with IFN‐γ plus LPS in the NO production. Ginsenoside‐Rh2 also effectively inhibited IFN‐γ‐induced NO production when the cells were treated with IFN‐γ 6 hr after the treatment with ginsenoside‐Rh2. Our findings suggest that this phenomenon might be caused by inhibition of priming signal such as IFN‐γ for the synergistic induction of NO synthesis.
Normalization of immune response and phosphoinositide fatty acid composition of peripheral blood lymphocytes in an alcoholic patient after alcohol abstinence
Authors: Stefanini G. F. , Castelli E. , Foschi F. G. , Marsigli L. , Addolorato G. , Celadon M. , Biagi P. L. , Bordoni A. , Gasbarrini G. , Hrelia S.
Published: July 2007

After 10 months of alcohol abstinence a malnourished alcoholic patient improved his nutritional status. The analysis of peripheral blood lymphocyte response to mitogenic stimulation with the antibody anti‐CD3 and of the fatty acid composition of the (poly)‐phosphoinositide fraction derived from lymphocytes revealed: 1) a similar [3H]‐thymidine uptake as in control (non‐drinker) subjects; 2) a similar relative molar content of the main fatty acids in the (poly)‐phosphoinositides as in control subjects. Alcohol abstinence can normalize both the parameters, which are greatly altered during alcohol abuse. This suggests a link between nutritional status and lymphocyte responsiveness via phosphoinositide fatty acid composition.
Affinity purification of mannose 6‐phosphate receptor proteins. Purification and partial characterisation of goat liver receptors
Authors: Yerramalla Udaya Lakshmi , Nadimpalli Siva Kumar
Published: July 2007

Mannose 6‐phosphate receptor (MPR) proteins designated as MPR 300 and MPR 46 have earlier been purified from some mammals on phosphomannan coupled to cyanogen bromide activated Sepharose. In a recent study, the goat liver MPR 300 has been directly purified using Sepharose‐divinylsulfone‐pentamannosyl phosphate matrix (Sivakumar N. 1996, J.Biochem. Biophys. methods, 31, 181‐184(1)). In the present report, we describe the preparation of another affinity matrix Sepharose‐divinylsulfone‐phosphomannan and its utility in purifying the MPR proteins from goat liver. While the MPR 300 from goat liver showed an electrophoretic mobility similar to other mammalian MPRs, the small receptor showed a molecular weight of 36 kDa. Antibodies raised against goat liver MPR 300 react specifically with the large receptor protein. Additionally affinity purified peptide specific antibody corresponding to amino‐acid residues 26‐42 (ADGCDFVCRSKPRNVPA) of the cytoplasmic tail of the human liver MPR 46 (Pohlmann et al, 1988. Proc. Natl. Acad. Sci. USA, 84, 5575‐5579 (2)) cross‐reacts with the purified small receptor.
Catalytic roles of lysines (K9, K27, K31) in the N‐terminal domain in human adenylate kinase by random site‐directed mutagenesis
Authors: Ayabe Takanori , Park Seung Kyu , Takenaka Hitoshi , Sumida Michihiro , Uesugi Seiichi , Takenaka Osamu , Hamada Minoru
Published: July 2007

To elucidate lysine residues in the N‐terminal domain of human cytosolic adenylate kinase (hAK1, EC 2.7.4.3), random site‐directed mutagenesis of K9, K27, and K31 residues was performed, and six mutants were analyzed by steady‐state kinetics. K9 residue may play an important role in catalysis by interacting with AMP2‐. K27 and K31 residues appear to play a functional role in catalysis by interacting with MgATP2‐. In human AK, the ε‐amino group in the side chain of these lysine residues would be essential for phosphoryl transfer between MgATP2‐ and AMP2‐ during transition state.
Cloning, expression and characterization of a blood group B active recombinant α‐D‐galactosidase from soybean (Glycine max)
Authors: Davis Melissa O. , Hata D. Jane , Johnson Scott A. , Walker John C. , Smith Daniel S.
Published: July 2007

A cDNA encoding soybean α‐D‐galactosidase [E.C. 3.2.1.22] was obtained by screening a soybean library with Phaseolus α‐D‐galactosidase cDNA. The Glycine max α‐D‐galactosidase cDNA is 1.75 kb long and contains untranslated 5′ and 3′ sequences. The deduced amino acid sequence of the soybean gene has a high degree of homology with other eucaryotic α‐D‐galactosidases. Recombinant α‐D‐galactosidase (rGal) was expressed in Pichia pastoris and purified by affinity chromatography. Purified rGal was homogeneous as judged by SDS‐PAGE analysis with the relative molecular mass under reducing conditions of 39.8, and under nonreducing conditions 38.0 kDa. The expressed protein contained the sequence NGLGHTPPMG at the N‐terminus, corresponding to the deduced amino acid sequence of the soybean gene. The relative native molecular mass by Sephacryl S‐200 chromatography was determined to be 33.1 kDa. The specific activity was 295.6 μmoles of PNP‐α‐D‐galactopyranoside hydrolyzed per mg pure rGal per min. rGal was highly specific for α‐D‐galactosyl residues. No detectable hemagglutinin or protease activity was present in the preparations. Furthermore, rGal was active against the blood group B antigen in native human erythrocyte cell suspension assays. The only detectable erythrocyte phenotypic change was loss of the B and P1 epitopes. Consequently, recombinant Glycine max α‐D‐galactosidase may have useful biotechnical applications in the potential mass production of universally transfusable type O erythrocytes by enzymatic conversion.
β1 integrin subunit dimerization via disulfide bonds
Authors: Belkin Vladimir M. , Kozlova Nadezhda I. , Bychkova Victoria V. , Shekhonin Boris V.
Published: July 2007

Integrins of the β1 family were isolated from human smooth muscle. SDS‐PAGE analysis and subsequent immunoblotting demonstrated that integrin samples contain a protein immunologically related to β1 integrin subunit with the previously undescribed apparent molecular mass 205 kD. One‐dimensional peptide mapping showed that the 205 kD protein is not a novel β1 related integrin subunit, but a β1 integrin subunit dimer. After reduction the major part of the β1 immunoreactive material migrated from the 205 kD to 130 kD region, indicating that β1 integrin subunit dimers were formed via disulfide bonds. When electrophoretically pure β1 monomer and dimer forms were analized it was found that during SDS‐PAGE about 30% of β1 integrin subunit monomers were organized into dimers while approximately 70% of the β1 dimer form was partly disrupted into monomers. It was suggested that this steady‐state process is a result of a reversible reaction between intra‐ and intermolecular disulfide bonds. Possible in vivo dimerization of integrins via disulfide bonds is discussed.
Characterization of natriuretic peptide receptors in the rat parotid
Authors: Nashida Tomoko , Imai Akane , Shimomura Hiromi
Published: July 2007

Competition studies between 125I‐ANP and ANP (atrial natriuretic peptide) or the ring‐deleted analog C‐ANF4‐23 (C‐ANF) revealed the presence of the A‐type natriuretic peptide receptor (GC‐A receptor) and the ANP‐clearance (ANP‐C) receptor in the rat parotid membrane. The ratio of the GC‐A and ANP‐C receptors was about 3 to 1. Enhancement of the cGMP level by the C‐type natriuretic peptide (CNP) revealed the presence of the B‐type natriuretic peptide receptor (GC‐B receptor) in the rat parotid gland. Based on the binding studies with ANP and C‐ANF and cGMP analysis by ANP and CNP, the natriuretic peptide receptors in the rat parotid gland were demonstrated to have a rank order on the amount of GC‐A receptor > GC‐B receptor > ANP‐C receptor.
Proton pump and auxin effect in Arabidopsis thaliana leaves during the development
Authors: Bellamine Jalil , Greppin Hubert
Published: July 2007

The plasma membrane from Arabidopsis thaliana leaves (wild‐type and F mutant) has been purified by two‐phase partitioning and the H+‐transport activity was tested in vitro in the presence of different concentrations of IAA. While 1 µM of IAA had no effect, higher concentrations (10 µM and 100 µM) were inhibitory in 25 short days grown wild‐type plants. However, the activity was increased by about 16 % after 10 supplementary short days of growth (from 25 to 35 SD) in the represence of 10 µM of IAA. No significant sensitivity to the tested concentrations of auxin was observed during the development in short days (30, 38, 42 SD) or even the plants were induced by a 24 h of continuous light. The variability was not observed for a given developmental stage, from one experiment to another, but it was also observed after light induction of F mutant plants.
Kinetics of inhibition of green crab (Scylla serrata) alkaline phosphatase by phenylglyoxal
Authors: Xie Wen‐Zhang , Wang Hong‐Rui , Chen Qing‐Xi , Zhou Hai‐Meng
Published: July 2007

The kinetics method of the substrate reaction during modification of enzyme activity previously described by Tsou (Adv. Enzymol. Related. Areas Mol. Biol., 1988, 61, 381‐436) has been applied to study the kinetics of the course of inactivation of green crab alkaline phosphatase by phenylglyoxal. The obtained results shows that the inactivation of the enzyme by phenylglyoxal is a slow reversible reaction. The microscopic rate constants for the reaction of the inhibitor with the enzyme were determined. The presence of substrate offers marked protection of this enzyme against inactivation by phenylglyoxal. The above results suggest that the arginine residue is essential for activity and is situated at the active site of the enzyme.
A novel DNA cleaving agent, 2,2′‐bis(2‐aminoethyl)‐4,4′‐bithiazole, induces thymocyte apoptosis
Authors: Lee Eibai , Miyaguchi Fumiko , Inoue Maho , Kariya Kimio , Sasaki Hideaki
Published: July 2007

The incubation of rat immature thymocytes with 2,2‐bis(2‐aminoethyl)‐4,4′‐bithiazole, which possesses the cleaving activity toward plasmid DNA in the presence of Co2+, resulted in an increase in DNA fragmentation. No DNA fragmentation of the cells was induced by 2,2‐bis(3‐aminopropyl)‐4,4′‐bithiazole, which has little cleaving activity toward plasmid DNA. Analysis of fragmented DNA from thymocytes treated with 2,2‐bis(2‐aminoethyl)‐4,4′‐bithiazole by agarose gel electrophoresis revealed that the compound produced 50 kbp DNA fragments and caused internucleosomal DNA fragmentation. In addition, the confocal microscopic images of thymocytes treated with the compound showed condensation of chromatin. These results indicate that bithiazole derivative induces thymocyte apoptosis with the biochemical and morphological characteristics through triggerring DNA scission.
Does an increase in dietary linoleic acid modify tissue concentrations of cervonic acid and consequently alter alpha‐linolenic requirements? Minimal requirement of linoleic acid in adult rats
Authors: Bourre J‐M. , Dumont O. , Durand G.
Published: July 2007

Rats were fed a control diet containing both linoleic and alpha‐linolenic acid. When 60‐days‐old they were divided into 8 groups, each receiving the same amount of alpha‐linolenic acid, but varying amounts of linoleic acid. When the (n‐6)/(n‐3) ratio in the diet varied from 2 to 32 (with a constant amount of 150 mg alpha‐linolenic acid per 100 g diet), tissue levels of the (n‐3) series fatty acids were not significantly modified, except in the liver, heart and testes. In all organs studied, the saturated and monounsaturated fatty acids were practically unchanged. For the (n‐6) series fatty acids, arachidonic acid was not significantly affected, in muscle, kidney, brain, myelin, nerve‐endings or sciatic nerve, whatever the quantity of linoleic acid in the diet. In liver, arachidonic acid plateaued at 2400 mg linoleic acid/ 100 g diet and at 400 mg /100g diet in heart. Results for 22:5(n‐6) showed a marked increase in heart, a moderate increase in liver and kidney, and no effect in muscle, testes, brain, myelin, nerve‐endings or sciatic nerve. This experiment defined the minimum amount of linoleic acid required in the diet to maintain fatty acids of the linoleic family in the young adult rat. For the first time it was demonstrated that 1200 mg/100 g diet are sufficient for the liver, as evidenced by maintenance of the arachidonic acid concentration. For the other organs, there is either a very marked preservation of this acid, or the dietary level is less than 300 mg/100 g diet. For the essential fatty acid precursors (i.e. linoleic and alpha‐linolenic acids), the optimal (n‐6)/(n‐3) ratio required in the diet is about 8.
tRNATrp as cofactor of gelonin, a ribosome‐inactivating protein with RNA‐N‐glycosidase activity features required for the cofactor activity
Authors: Brigotti Maurizio , Carnicelli Domenica , Alvergna Paola , Pallanca Alessandra , Sperti Simonetta , Montanaro Lucio
Published: July 2007

Purified beef and rabbit liver tRNATrp, but not yeast tRNATrp, increase the in vitro inactivation of eukaryotic ribosomes by gelonin, a ribosome‐inactivating protein with RNA‐N‐glycosidase activity on 28S rRNA. Aminoacylation and stepwise trimming of the 3′‐end of bovine tRNATrp affect the cofactor activity, the most active species being that shortened by the last two nucleotides. ATP only moderately increases the activity of purified mammalian tRNATrp and in this increase the cognate aminoacyl‐tRNA synthetase apparently has no role. Mobility shift experiments indicate that bovine tRNATrp binds both to ribosomes and to gelonin and favours the dissociation of gelonin from ribosomes.
Epitope mapping of preS2 of the hepatitis B virus surface antigen against a conformation‐dependent monoclonal antibody using synthetic peptides
Authors: Lee Myung Kyu , Kim Kil Lyong , Hahm Kyung‐Soo
Published: July 2007

Previously we reported that the N‐terminal sequence 120/123‐129 of the preS2 region of the hepatitis B virus surface antigen plays an important role on peptide antigenicity against a monoclonal antibody H8 (H8 mAb) by affecting the B cell epitope conformation of a peptide existing within the sequence 130‐145 (Lee et al., Biochem. Mol. Biol. Int., 34, 159‐168, 1994). In this study, we try to map the H8 mAb binding site using a series of substituted peptides in the sequence 131‐143 by competitive ELISA. Peptide antigenicities were greatly reduced when the residues 131 (L), 137 (R), 140 (Y), 141 (F) and 142 (P) were substituted. The residues 133 (D), 134 (P) and 136 (V) had a slight affect on the mAb binding, whereas the residues 135 (R) and 139 (L) had no effects on the mAb binding. In contrast to H8 mAb, however, three anti‐HBsAg polyclonal antisera showed the lowest bindings to the peptide substituted at position 135. These results suggest that the epitope against H8 mAb is discontinuously conformational.
Modulation of ouabain sensitive sodium potassium pump of erythrocytes from patients with chronic renal failure:Role of acute hemodialysis
Authors: Prasad R. , Mond R. , Jain S. , Kaur G. , Chugh K. S.
Published: July 2007

Significantly higher levels of plasma urea creatinine and potassium were observed in patients with renal failure compared to normal controls. The RBC sodium concentration was raised whereas the RBC potassium concentration was decreased in chronic renal failure. These alterations in the RBC Na1 and K1 concentrations were associated with decrease in ouabain sensitive sodium efflux rate and ouabain sensitive sodium efflux rate constant. However, there was no significant impact of acute hemodialysis on the intracellular electrolytes levels, ouabain sensitive sodium efflux rate and ouabain sensitive sodium efflux rate constant. These findings suggest an intrinsic alteration in the transport capacity of Na+‐K+ pump which could account for the rise in intracellular sodium and fall in intracellular potassium content in the RBCs of chronic renal failure patients.
The effects of decyl aurachins C and D on the respiratory electron flow of facultative phototrophic bacteria
Authors: Romagnoli Simona , Oettmeier Walter , Zannoni Davide
Published: July 2007

Decyl aurachins C and D are two synthetic compounds related to the natural products aurachins C and D extracted from Stigmatella aurantiaca. Titrations of a range of partial respiratory activities in membranes from facultative phototrophs indicate that decyl aurachin C is more effective than aurachin D in inhibiting the quinol oxidase. Decyl aurachin C also affects the NADH‐ubiquinone oxidoreductase of Rhodobacter capsulatus and the cytochrome bc1 complexes of Rb. capsulatus and Rhodospirillum centenum. Titration of the light‐induced respiration in Rsp. centenum allowed us to estimate an upper limit for the ubiquinol oxidase concentration of 1.5±0.2 nmol/mg protein, or a ubiquinol oxidase/cytochrome c oxidase ratio of 3:1.
Antioxidant property of Fructus Momordicae extract
Authors: Shi Honglian , Hiramatsu Midori , Komatsu Makiko , Kayama Takamasa
Published: July 2007

The antioxidant effect of Fructus Momordicae extract, FME (mogrosides 75‐80%), was studied. FME reduced the stable free radical 1,1‐diphenyl‐2‐picrylhydrazyl (DPPH) and scavenged superoxide radicals (O2‐) generated by a hypoxanthine and xanthine oxidase system. It also scavenged hydroxyl radicals (⊎OH) generated by Fenton reaction. In addition, FME inhibited Fe(II) induced lipid peroxidation in rat cortex homogenates in a dose‐dependent manner, as indicated by decreased thiobarbituric acid‐reactive substances (TBARS) formation. Oral administration of FME inhibited TBARS and malonaldehyde (MDA) formation in the ipsilateral cortex 30 min after iron‐salt injection into the left cortex of rat. FME showed inhibitory effect on 4‐hydroxy‐2(E)‐nonenal (4‐HNE) formation induced by Fe(III) injection into the rat cortex. These data suggest that Fructus Momordicae extract has an antioxidant activity against free radicals and lipid peroxidation.
Chlorpromazine transport in membrane vesicles from multidrug resistant CCRF‐CEM cells
Authors: Syed Samiuddin K. , Christopherson Richard I. , Roufogalis Basil D.
Published: July 2007

The mechanism by which chlorpromazine (2‐chloro‐10‐(3‐dimethylaminopropyl)‐phenothiazine) reverses P‐glycoprotein (P‐gp2)mediated multidrug resistance was investigated using membrane vesicles prepared from human CCRF‐CEM leukaemia cells. Chlorpromazine was transported in an ATP‐dependent manner into membrane vesicles prepared from vinblastine resistant (VBL1000) cells but not from drug‐sensitive cells. The chlorpromazine uptake was sensitive to osmotic pressure, indicating true transport into the vesicle lumen. The ATP‐dependent chlorpromazine uptake was inhibited about 30% by the addition of ammonium chloride, indicating that a pH or electrochemical gradient could not account for the majority of ATP‐dependent chlorpromazine uptake. The results of this study show that chlorpromazine is actively transported by P‐glycoprotein and that chemosensitization by phenothiazines may occur by competition of these agents for active transport of anticancer agents by P‐glycoprotein.
Changes in fatty acid composition in rat adipose tissue induced by dietary obesity
Authors: Lladó Isabel , Pons Antoni , Palou Andreu
Published: July 2007

The aim of the study was to evaluate the effects of cafeteria feeding on the composition of fatty acids in retroperitoneal fat pad and also to determine what happens to fatty acids when rats previously fed the cafeteria diet are returned to regular rat chow. The study of the post‐cafeteria rats enabled us to determine the effects of dietary induced excess weight in the absence of artefactual interferences from the diet because these rats, unlike the cafeteria obese rats, ate the same diet as controls.

In response to cafeteria feeding there were increases in the majority of adipose tissue fatty acids. However, significant decreases were observed in the relative proportions of 18:2n‐6 and in two related n‐6 polyunsaturated fatty acids (18:3n‐6 and 20:3n‐6), as well as in 16:1. In the post‐cafeteria obesity model the previous dietary influence on fatty acid composition was still evident. The maintenance of both the high levels and proportions of 18:1 and the decrease of 16:1 percentage in the post‐cafeteria rats argues in favour of an alteration in the activity of the elongation metabolic pathway. Certain changes affecting polyunsaturated fatty acid adipose depot composition of obese rats, mainly the decreased levels of 18:2n‐6, are long lasting and could be related to the maintenance of the obese status. On the whole, although the fatty acid composition of adipose tissue is influenced by the composition of the diet, there are some differences in both the maintenance of the effects and also in the selectivity of adipose tissue for the different fatty acids of obese and lean rats.
Rapid assessment of early glycation products by mass spectrometry
Authors: Prabhakaram Malladi , Smith J. B. , Ortwerth B. J.
Published: July 2007

In order to detect the early glycation products, we have reacted a model peptide (t‐boc‐lys‐ala‐ala) with L‐threose (a degradation product of ascorbic acid) and analyzed the reaction products by a combination of HPLC and mass spectrometry. Amino group modification, as observed by a fluorescamine assay, indicated complete modification after 3 days of incubation with a 10‐fold excess of threose. As much as 60% of the adducts were acid labile and only 4% of the adducts could be observed by amino acid analysis. However, Fast atom bombardment mass spectrometry (FABMS) of the samples incubated for 6 hr showed relative molecular masses consistent with the formation of adducts corresponding to the addition of one and two molecules of L‐threose to the peptide. Likewise, samples incubated for 12 hr showed peptide adducts with two and three L‐threoses. The number of threose molecules added to the peptide was also confirmed from the FABMS analysis by using [1‐13C]‐threose as the glycating agent.
1H‐NMR relaxometric study of pancreatic serine (pro)enzyme inhibition by a Gd(III) chelate bearing boronic functionalities
Authors: Aime Silvio , Fasano Mauro , Paoletti Silvia , Viola Franca , Tarricone Cataldo , Ascenzi Paolo
Published: July 2007

Binding of the paramagnetic N,N″‐bis(m‐boroxyphenylcarbamoylmethyl)‐diethylenetriamine‐N,N′,N″‐triacetic acid Gd(III) complex (GdBB) to chymotrypsin, chymotrypsinogen, trypsin, trypsinogen and pancreatic elastase has been investigated by 1H‐NMR relaxometry, between pH 6.0 and 8.5, at 25.0°C. Values of Ki for the competitive inhibition of serine proteinases by GdBB are in excellent agreement with values of Kd obtained by 1H‐NMR relaxometry, suggesting that the substrate and the paramagnetic complex bind to the same region. Moreover, 1H‐NMR relaxometry allowed to determine values of Kd for GdBB binding to chymotrypsinogen and trypsinogen, both devoid of catalytic activity. The increase of the water proton relaxation rate upon GdBB binding to serine (pro)enzymes may be useful in the design of novel functional contrast agents for magnetic resonance imaging.
A novel cDNA encoding for a lim domain protein located at human chromosome 14q32 as a candidate for leukemic translocation
Authors: Tsui Stephen Kwok Wing , Chan Peggy Pik Kwan , Cheuk Chin Wah , Liew Choong Chin , Waye Mary Miu Yee , Fung Kwok Pui , Lee Cheuk Yu
Published: July 2007

A full‐length cDNA clone encoding a zinc finger protein was isolated and sequenced. This full‐length clone consists of 728 bp and has a predicted open reading frame (ORF) encoding 208 amino acids. The ORF of this polypeptide codes for the human cysteine‐rich protein 2 (HCRP2) and has an amino acid sequence that is 92.8% identical to its rat homolog (RCRP2). HCRP2 was mapped to chromosome 14q32, which is a hot spot of translocation in tumor development, by fluorescent in situ hybridization (FISH).
3‐Hydroxybutyrate decreases noradrenaline affinity for rat erythrocyte ghost β2‐adrenergic receptors
Authors: Gómez G. , Cañas X. , Fernández‐López J. A. , Remesar X. , Alemany M.
Published: July 2007

The effects of physiological levels of 3‐hydroxybutyrate on noradrenaline binding to rat erythrocyte membranes were studied. The binding of noradrenaline to membranes was determined by measuring the union of 3H‐noradrenaline in the presence of varying concentrations of 3‐hydroxybutyrate. Scatchard plots of the results were used to determine that 3‐hydroxybutyrate provoked a loss of affinity of the β2‐adrenergic receptors for adrenaline. Thus high (but physiological) 3‐hydroxybutyrate levels can lower the binding of the catecholamine, thus diminishing the extent or intensity of adrenergic‐driven responses
Characterization of the purified cytosolic thymidine kinase from murine Ehrlich ascites tumor: Interconversion of two different relative molecular weight forms
Authors: Tsukifuji‐Nabeya Reiko , Yusa Toshikazu , Kuroiwa Namiko , Kumazawa Takeshi , Tamiya Norihisa , Moriyama Yoichi , Okamoto Shoji , Yamaguchi Yutaka , Fujimura Shinji
Published: July 2007

Cytosolic thymidine kinase was purified 18,000‐fold of the homogenate from murine Ehrlich ascites tumor, using the [p‐aminophenyl 3′‐dTMP]‐CH‐Sepharose affinity column. Sodium dodecyl sulfate‐polyacrylamide gel electrophoresis of the purified enzyme showed a single protein band of molecular weight 26,000. Two different forms, relative molecular weight 50,000 and 70,000, were found by gel filtration, depending on the existence of dithiothreitol, ATP and other nucleotides. These agents also stabilize and stimulate the enzyme activity. The existence of two forms was also manifested by DEAE‐Sephacel column chromatography, where the 50,000 form was eluted by 50 mM NaCl and the 70,000 form by 400 mM NaCl.
Effect of a cholesterol‐rich diet on the excitability of rabbit aorta
Authors: Wong Kai K.
Published: July 2007

The level of plasma total cholesterol in rabbits after 8‐week feeding on a cholesterol‐rich diet was raised significantly. The excitability of the rabbit aorta in response to the depolarizing agent KC1 was enhanced, demonstrating an increased sensitivity to KC1 stimulation as well as an increase of the KC1‐induced maximum tension. It is speculated that 7 weeks of hypercholesterol state may vary the membrane lipid composition, enhancing the aortic excitability in response to depolarizing agent.
A low molecular weight urokinase derivative with enhanced fibrin affinity
Authors: Chen Xiao‐Chun , Hua Zi‐Chun , Zhu De‐Xu
Published: July 2007

A Gly‐Pro‐Arg‐Pro tetrapeptide, homologous to amino‐terminal segment of the human fibrin α chain after the release of the fibrinopeptide A, was covalently coupled to peptide A of low molecular weight urokinase. The resulting derivative gained increased affinity for fibrin. In caseinolytic assay, fibrin can stimulate the derivative to activate plasminogen. The derivative had two‐fold greater fibrinolytic potency than native low molecular weight urokinase and its affinity for fibrin clot was 3.9‐fold higher than that of low molecular weight urokinase.
Binding of non‐homologous series of anticancer drugs to cytosine
Authors: Cserháti Tibor , Forgács Esther
Published: July 2007

The interaction of 20 anticancer drugs with cytosine was studied by charge‐transfer reversed‐phase thin‐layer chromatography. The hydrophobicity, the specific hydrophobic surface area and the relative strength of interaction was calculated. Significant linear correlation was found between the hydrophobicity and specific hydrophobic surface area of drugs; this means that, from the chromatographic point of view, they form a homologous series of solutes. Only eight anticancer drugs showed significant interaction with cytosine. The binding of these drugs to cytosine (and possibly to other nucleotides) might be an important interaction which plays a role in the biological activity of drugs.
Hysteretic properties of rabbit skeletal muscle phosphorylase kinase: Synergistic activation by phosphorylase b, Ca2, and Mg2
Authors: Kurganov B. I. , Andreeva I. E. , Makeeva V. F. , Livanova N. B.
Published: July 2007

To study the hysteretic properties of rabbit skeletal muscle phosphorylase kinase the method of continuous registration of the kinetics of the kinase reaction developed by us earlier has been used. It was shown that duration of the lag period on the kinetic curves is independent of the phosphorylase kinase concentration and the simultaneous presence of phosphorylase b, Ca2+, and Mg2+ is required for the complete transition of the enzyme into the activated state.
cDNA sequence analysis of a novel cardiotoxin‐like protein from Taiwan banded krait
Authors: Chang Long‐Sen , Lin Jordge
Published: July 2007

The cDNAs encoding a novel protein was constructed from the cellular RNA isolated from the venom glands of Bungarus multicinctus by reverse transcription‐polymerase chain reaction. The deduced amino acid sequence of this novel protein contained 65 amino acid residues with 8 cysteine residues. Comparative sequence analysis showed that it was structurally related to cardiotoxin rather than neurotoxins. These results suggest that the venom of Bungarus multicinctus may contain cardiotoxin(s) which was not noticed before.
Intracellular distribution of polyamines in human lymphoblastoid cell line during phorbol ester‐induced differentiation
Authors: Stabellini Giordano , Creati Beatrice , Primio Roberto Di , Trubiani Oriana
Published: July 2007

Polyamines such as putrescine, spermidine and spermine play an important role in nucleic acid metabolism. These aliphatic amines display a key role in cell‐induced transformation by carcinogenic substances. In particular, one of these, the phorbol myristate acetate, provokes cell differentiation and gives an increase of ornithindecarboxylase activity; enzyme regulating the pathways of polyamines. In this study we analyse the trend of the polyamines at cytoplasmic and nuclear level during phorbol treatment. Our results show a correlation between nuclear and cytoplasmic spermine, 3H‐Thymidine, 3H‐Leucine incorporation and cell cycle phases. These data remark that the polyamines are differently distributed into the cell during the phorbol myristate acetate‐mediated differentiation process and that the spermine is down‐regulated for to supply the increased protein biosynthesis.
Sequence‐specific interactions of minor groove binders with the 154 base pair hindlll‐rsal restriction fragment of cDNA of the human tau 40 protein involved in pathology of alzheimer's disease
Authors: Kittler Leonhard , Matesoi Diana , Bell Achim , Baguley Bruce C. , Unger Eberhard , Löber Günter
Published: July 2007

The DNA minor groove binders netropsin, distamycin and four structurally related bisquaternary ammonium heterocycles (BQA), SN 6999, SN 6570, SN 6132 and SN 6131, were investigated for sequence‐specific interactions with the 154 base pair fragment of cDNA of the human Tau 40 protein (h Tau 40 protein), involved in pathology of Alzheimer's disease. The base sequences 5′ AATCTT 3′, 5′ AATATT 3′ and 5′ TTTCAATCTTTTTATTT 3′ were identified as ligand specific binding sites and demonstrate the obvious dA·dT binding preference. Footprinting titration experiments were performed to estimate sequence‐specific binding constants (KA). The KA‐Values were in the order of 106M‐1 and dependent on DNA base sequence as well as ligands used. The highest values estimated were for netropsin (KA=5.0×106M‐1) and the quinoline derivative SN 6999 (KA=6.2×106M‐1) binding to the sequence 5′ ATAAT 3′. Microscopic binding constants are determined by the base sequence rather than by the length of dA·dT stretches. In the extended dA·dT run, 5′ TTTCAATCTTTTTATTT 3′, netropsin and distamycin binding tolerates the presence of two dG·dC base pairs, as indicated by nearly unaffected footprints. In contrast, the failure of BQAs to form footprints demonstrates their significantly decreased binding selectivity.
Cell volume regulation in Claviceps fusiformis. An animal‐type Na,K‐ATPase operating in a fungus?
Authors: Rauferová Ludmila , Metlička Rudolf , Beneš Ivan , Kotyk Arnošt , Janáček Karel
Published: July 2007

Claviceps fusiformis, an ergot pathogen of pearl millet, was equilibrated in media without and with 0.1 mM ouabain (g‐strophantin). Highly significant effects of ouabain on the contents of sodium ions and water were found, suggesting that an active transport of sodium ions, similar to that in animal cells, regulates the cell volume of the fungus. The basic role of the sodium pump appears to be to decrease larger cell volumes and to increase the small ones; for this purpose it can even reverse its direction.
Effect of intramitochondrial Mg2 on citrulline synthesis in rat liver mitochondria
Authors: Rodríguez‐Zavala José S. , Saavedra‐Molina Alfredo , Moreno‐Sánchez Rafael
Published: July 2007

A correlation was found between mitochondrial matrix free magnesium ([Mg2+]m), measured by means of the fluorescent indicator Mag‐Fura‐2, and citrulline synthesis in rat liver mitochondria. The variation of [Mg2+]m from 0.05 to 1.7 mM by changes in external Mg2+, induced a 20 + 8.5% increase in the rate of citrulline synthesis, whereas a further increase of [Mg2+]m to 3.3 mM induced a return to basal values. The increase in [Mg2+]m, as well as the diminution of external pH, also promoted an elevation of matrix free Ca2+ ([Ca2+]m). An increase in [Ca2+]m, at constant [Mg2+]m and pH, resulted in a 3‐fold stimulation of citrulline synthesis. The data suggest that [Mg2+]m may modulate the rate of citrulline synthesis through a direct interaction with carbamoyl‐phosphate synthase I (ammonia) and, indirectly. by changing the levels of matrix Ca2+.
Interaction of recombinant rat nucleoside diphosphate kinase α with bleached bovine retinal rod outer segment membranes: A possible mode of pH and salt effects
Authors: Orlov Nicolay Ya. , Orlova Tatiana G. , Reshetnyak Yana K. , Burstein Edward A. , Kimura Narimichi
Published: July 2007

An attempt was made to reveal the mode of action of protons and salts on the recently discovered GTPγS‐dependent interaction of bovine retinal rod outer segments (ROS)1 nucleoside diphosphate kinase (NDP kinase) with the complex between bleached visual receptor rhodopsin and retinal G‐protein transducin in bovine ROS membranes. The properties of recombinant rat NDP kinase α, that is immunologically similar to the soluble NDP kinase from bovine ROS preparation, have been studied in solution by means of protein fluorescence at different pH and salt concentrations and results were compared with pH and salt effects on the binding of NDP kinase α to bleached bovine ROS membranes. The results suggest that NDP kinase α itself may serve as a target for protons and salts and mediates their effects on the interaction between the enzyme and ROS membranes.
Identification of the functional improtance of valine‐19 residue in streptokinase by N‐terminal deletion and site‐directed mutagenesis
Authors: Lee Si Hyoung , Jeong Seong Tae , Kim Il Chul , Byun Si Myung
Published: July 2007

Streptokinase (SK) is a bacterial plasminogen activator of multi‐domain structure. In deletion analysis of the N‐terminal region of SK, the deletion of 20 amino acids (SKΔN20) resulted in the dramatic reduction of plasminogen activator activity compared to deletion of 7 (SK ΔN7) and 13 amino acids (SKΔN13). The incubation time to reach maximal active site generation in an equimolar mixture of SKΔN20 and plasminogen was the same as that for wild‐type SK. To identify the functional residues important in plasminogen activation, several site‐directed mutations were introduced at the region spanning Serl6‐Val20 of SK. The results showed that Vall9 residue is important for the activity of the SK‐plasminogen complex.
Immunoreactivities of p11 and calcyclin in baker's yeast, wheat germ and lobster
Authors: Kuo W. N. , Jn‐Baptiste J. B. , Zhai L. , Chambers M. C. , Kanadia R.
Published: July 2007

After effectively eliminating the nonspecific cross‐immunoreactivity with the affinity columns of anti‐IgG agarose and IgG agarose, the potent immunoreactivities of p11 and calcyclin in wheat germ, lobster tail muscle, and three strains of baker's yeast were analyzed by Western blotting using mouse anti‐p11 and rabbit anti‐calcyclin. The occurrence of multiple bands may be due to either autolyses and/or the interactions between the p11 (or calcyclin) and other endogenous biological molecules. The results suggest not only a ubiquitous distribution and a universal Ca2+‐mediating regulatory role of p11 and calcyclin in eukaryotes, but also an evolutionary conservation of these (S‐100)‐related proteins.
Opposite effects of isoniazid and fasting on the expression of CYP2E1 protein and mRNA in house musk shrew (Suncus murinus)
Authors: Nakura Hironori , Itoh Susumu , Kusano Hisashi , Ishizone Hiroyuki , Kamataki Tetsuya
Published: July 2007

The effects of isoniazid and fasting on hepatic CYP2E1 in the suncus were investigated. Aniline hydroxylation and N‐nitrosodimethylamine demethylation, which are known to be catalyzed by CYP2E1, in liver microsomes were induced and suppressed by the treatment with isoniazid and the fasting, respectively. Immunoblot analysis indicated that CYP2E1 protein in liver microsomes from isoniazid‐treated suncus was increased in contrast to the result with the fasting of the suncus. Northern blot analysis showed that the treatment of suncus with isoniazid increased the expression of CYP2E1 mRNA in livers although the fasting of the suncus significantly decreased CYP2E1 mRNA. These results suggest that the regulation of hepatic CYP2E1 in the suncus by treatment with isoniazid and fasting was different from that in rats.
Inhibition of DNA synthesis in CCL 39 cells by impermeable iron chelators
Authors: Alcaín Francisco J. , Löw Hans , Crane Frederick L.
Published: July 2007

The synthesis of DNA in CC1 39 cells is inhibited by the presence of the Fe2+ chelator bathophenanthroline disulfonate (BPS) when growth is stimulated by thrombin EGF plus insulin, but not by fetal calf serum. The presence of transferrin and Fe3+ in fetal calf serum can be the basis for lack of BPS effect with serum. The impermeable Fe3+ chelator Tiron does not, by itself inhibit growth factor induced DNA synthesis, but it induces together with BPS inhibition on fetal calf serum induced DNA synthesis. The combined effect of BPS and Tiron is similar to inhibition of DNA synthesis by impermeable polyvalent DTPA which can chelate both Fe2+ and Fe3+ but does not inhibit ribonucleotide reductase in intact cells. Ferrous iron that bind BPS can relieve the inhibition at stoichiometric concentration. Ferric iron also prevents the inhibition even though it does not bind BPS. BPS does not inhibit DNA synthesis in HeLa cells. BPS reacts with iron from CC1 39 cells but not from HeLa cells. Data show that iron available for impermeable external chelators is in the ferrous state, and that exogenous iron should be reduced before it reverses the inhibition.
Changes in protein composition during chick embryo retina development
Authors: Carabillò M. , Gerbino E. , Main H. , Calvaruso G. , Tesoriere G.
Published: July 2007

Two dimensional gel electrophoresis was employed to analyse the protein composition of chick embryo retina during “in ovo” development in order to individuate some components which are typical of different stages of growth. The study of protein composition of 7‐day‐old chick embryo retinas revealed the presence of about eighty different soluble components. In four of these proteins the staining intensity decreased during retina development. One of these components, of 35 KD with an isoelectric point of about 5.15, was partially purified by means of preparative isoelectric focusing. Other components, which were absent at day 7 appeared during the second week of development and were clearly visualised at day 16. These results were also compared with those obtained by means of 2DGE performed on liver and brain of chick embryo at day 7 and 16 of development.
Purification and partial characterization of chromogranin‐A from human brain
Authors: Kumar Ujendra
Published: July 2007

Chromogranin A was purified from post‐mortem human brain tissue, obtained at autopsy by a three step chromatography procedure to a single band purity on SDS gel electrophoresis at 68 kDa. Purified protein was further characterized by immunoblotting using a mouse anti‐chromogranin A monoclonal antibody LK2H10, which recognizes the 68 kDa chromogranin A in human brain. The results demonstrate a simple protocol of preparing CgA from the human brain. Chromogranin plays a very important role in various neurological disorder and has been examined immunohistochemically in Alzheimer's disease, Parkinson's disease, Pick's disease. The procedure of purification may be useful in further defining its biological function, biochemistry and physiological significance in the central nervous system and may be useful in delineating the molecular pathology of certain neurological disorders.
The effects of chronic administration of nimodipine on age‐related changes in nitric changes in nitric oxide and its synthase in senescence‐accelerated mouse brain
Authors: Inada Kazunori , Yokoi Isao , Kabuto Hideaki , Namba Yukiko , Ogawa Norio
Published: July 2007

The levels of nitric oxide (NO) and NO synthase (NOS) activities in the brain of young‐adult (3 months old), aged (11 moths old) and nimodipine‐administered (11 months old) senescence‐accelerated mouse (SAM), of which SAMP8 sub‐strain is inferior in acquisition of learning and has a lower content in testosterone, were compared. Nimodipine, which is L‐type calcium ion channel blocker and has memory‐enhancing effects, was administered orally for 5 months. In the cerebral cortex of aged SAMP8, NOS activity was increased compared with that of young‐adult SAMP8. Though nimodipine did not alter the contents of NO in any brain regions compared with those in aged SAMP8, nimodipine increased NOS activity in the aged cerebellum. Our data suggest that nimodipine may increase NOS activity through elevation of testosterone level, as testosterone increases NOS only in the cerebellum, although further work is clearly needed to ascertain effects of nimodipine on testosterone metabolism and maintenance in the acquisition of learning.
Protective effect of vitamin E against ethionine toxicity
Authors: Padma P. , Setty O. H.
Published: July 2007

The effect of administration of ethionine on rat liver mitochondrial functions and the protective effect of vitamin E on ethionine induced damage was studied. Ethionine treatment decreased the rate of respiration, respiratory control ratio and P/O ratio. There was a significant decrease in the activities of NADH dehydrogenase, succinate cytochrome C reductase and cytochrome oxidase. A significant decrease was seen on membrane potential and on the levels of ATP. Among the mitochondrial phospholipids only cardiolipin decreased significantly. The lipid peroxide level increased significantly in ethionine treated rats. Administration of vitamin E prior to ethionine treatment relieved the effects (induced by ethionine) on all the parameters studied. This study shows that vitamin E protects against ethionine toxicity.