bluuebunny/crossref_metadata_2025_split huggingface dataset:
Isolation and characterization of a tumor‐derived human protein related to chromogranin A and its in vitro conversion to human pancreastatin‐48
Authors: TAMAMURA Hirokazu , OHTA Mitsuhiro , YOSHIZAWA Kumi , ONO Yoshiyuki , FUNAKOSHI Akihiro , MIYASAKA Kyoko , TATEISHI Kayoko , JIMI Atsuo , YAJIMA Haruaki , FUJII Nobutaka , FUNAKOSHI Susumu 
Published: March 2005

A protein with pancreastatin‐like immunoreactivity has been isolated and purified from liver metastasis of a patient with insulinoma. NH2‐terminal residue analysis, in conjunction with the use of antibodies that are specific for the C‐terminal amide peptide of porcine pancreastatin, identified this protein as a 186‐amino‐acid protein corresponding to human chromogranin A‐116‐301 (the fragment corresponding to the positions from 116 to 301 of human chromogranin A). Digestion of this protein with trypsin yielded a 48‐amino‐acid peptide with the retention of full pancreastatin activity. Serum from patient with insulinoma contains a peptide specie(s) that comigrates with the 48‐amino‐acid pancreastatin, suggesting that this peptide might be a physiologically important circulation form of pancreastatin in humans.A sensitive radioimmunoassay was established using antibody developed against a synthetic 29‐amino‐acid peptide amide of pancreastatin. Immunocytochemical staining revealed that a major population of human pancreatic islet cells were immunoreactive to the antiserum but with varying intensity of staining. Pancreastatin‐like immunoreactivity was not observed in exocrine acinar cells.
Valyl‐tRNA synthetase from yeast
Authors: FREIST Wolfgang , CRAMER Friedrich 
Published: March 2005

For discrimination between valine and the 19 naturally occurring noncognate amino acids, as well as between valine and 2‐amino‐isobutyric acid by valyl‐tRNA synthetase from baker's yeast, discrimination factors (D) have been determined from kcat and Km values in aminoacylation of tRNAVal‐C‐C‐A. The lowest values were found for Trp, Ser, Cys, Lys, Mct and Thr (D = 90–870), showing that valine is 90–870 times more frequently attached to tRNAVal‐C‐C‐A than the noncognate amino acids at the same amino acid concentrations. The other amino acids exhibit D values between 1100 and 6200. Generally, valyl‐tRNA synthetase is considerably less specific than isoleucyl‐tRNA synthetase, but this may be partly compensated in the cell by valine concentrations higher than those of noncognate acids.In aminoacylation of tRNAVal‐C‐C‐A(3′NH2 discrimination factors D1 are in the range of 40–1260. From D values and AMP formation stoichiometry, pretransfer proof‐reading factors II1 were determined; post‐transfer proof‐reading factors II2 were determined from D values and AMP formation stoichiometry in acylation of tRNAVal‐C‐C‐A.II1 values (7–168) show that pretransfer proof‐reading is the main correction step, posttransfer proof‐reading (II2=1–7) is less effective and in some cases negligible.Initial discrimination factors were calculated from discrimination and proof‐reading factors according to a two‐step binding process. These factors, due to different Gibbs free energies of binding can be related to hydrophobic interaction forces, and a hypothetical ‘stopper’ model of the amino‐acid‐binding site is discussed.
Interdependence of tumor necrosis factor, prostaglandin E2, and protein synthesis in lipopolysaccharide‐exposed rat Kupffer cells
Authors: PETERS Thomas , KARCK Ulrich , DECKER Karl 
Published: March 2005

Kupffer cells are the main producers of tumor necrosis factor‐α (TNF; cachectin) and eicosanoids in the liver exposed to lipopolysaccharide (endotoxin; LPS). A very rapid but transient release of TNF is followed by a slow, steady synthesis of prostaglandin E2 (PGE2). TNF itself is able to provoke eicosanoid synthesis in Kupffer cells; the rate and pattern of prostaglandin production are similar to those observed after treatment with LPS. Anti‐TNF antibodies completely neutralize TNF action on Kupffer cells, thus ruling out any participation of contaminating LPS. LPS stimulation of PGE2 production in Kupffer cells is reduced by the antiserum to 50%, indicating an involvement of TNF in the stimulatory action of LPS. On the other hand, PGE2, a potent inhibitor of LPS‐elicited TNF release, is able to suppress LPS‐ but not TNF‐stimulated eicosanoid synthesis in rat Kupffer cells.In addition to this autocrine circuit, extrahepatic factors participate in the regulation of Kupffer cell activation: glucocorticoids not only inhibit TNF or prostaglandin production, they also reverse the LPS‐specific changes in the prostaglandin pattern of Kupffer cells.LPS, TNF or cycloheximide when given alone in the concentration range applied in this study do not affect the viability of rat Kupffer cells. However, the combinations of cycloheximide and either LPS or TNF cause rapid death of the cultured cells. The cytolytic potential of either combination cannot be alleviated by treatment with glucocorticoids.
The 1H‐NMR assignments of the aromatic resonances in complexes of Lactobacillus casei dihydrofolate reductase and the origins of their chemical shifts
Authors: BIRDSALL Berry , ARNOLD John R. P. , JIMENEZ‐BARBERO Jesus , FRENKIEL Thomas A. , BAUER Christopher J. , TENDLER Saul J. B. , CARR Mark D. , THOMAS Janette A. , ROBERTS Gordon C. K. , FEENEY James 
Published: March 2005

All the aromatic proton resonances in the 500‐MHzNMR spectra of Lactobacillus casei dihydrofolate reductase have been assigned for several of its complexes with inhibitors. For the complexes with methotrexate and trimethoprim this was achieved by using a combination of NMR techniques in conjunction with a selectively deuterated protein designed to simplify the spectra such that nuclear Overhauser effect (NOE) connections could be detected with greater ease and certainty. By correlating these NOE data with crystal structure data on related complexes it was possible to assign all the aromatic resonances and to extend these assignments to spectra of other complexes of dihydrofolate reductase.The conformation‐dependent chemical shifts observed for many of the resonances could be explained qualita‐tively, but not quantitatively, in terms of ring‐current shifts. The discrepancies between calculated ring‐current shifts and the observed conformation‐dependent shifts could not in general be accounted for satisfactorily in terms of carbonyl‐group anisotropic shielding contributions calculated using presently available models. In the case of the Hδ1,δ2 protons of Phe30 some of the discrepancy probably results from a difference in the conformation of the Phe ring between the solution and crystal states.
Fructose‐1,6‐bisphosphate‐activated pyruvate kinase from Escherichia coli
Authors: SPERANZA Maria L. , VALENTINI Giovanna , MALCOVATI Massimo 
Published: March 2005

The allosteric properties of the fructose‐1,6‐bis‐phosphate‐activated pyruvate kinase from Escherichia coli were examined in the presence of a number of fructose bisphosphate analogues, as well as of increased ionic strength (NaCl) and of the hydrogen‐bond‐breaking agent, formamide.Fructose 2,6‐bisphosphate, ribulose 1,5‐bisphosphate and 5‐phosphorylribose 1‐pyrophosphate gave allosteric activation (additive to that of fructose 1,6‐bisphosphate).Formamide always decreased Vmax, but left unchanged the Km for phosphoenopyruvate, while it decreased the concentration of fructose bisphosphate required to give half‐maximal activity (K0.5).NaCl increased the K0.5 for both phosphoenolpyruvate and fructose bisphosphate, leaving Vmax unchanged.These results are consistent with ionic binding of fructose bisphosphate through phosphates and with a critical role of hydrogen bonds in stabilizing both the inactive and the active enzyme conformers.
Complete amino‐acid sequence of glyceraldehyde‐3‐phosphate dehydrogenase from the hyperthermophilic eubacterium Thermotoga maritima
Authors: SCHULTES Verena , DEUTZMANN Rainer , JAENICKE Rainer 
Published: March 2005

The complete amino‐acid sequence of D‐glyceraldehyde‐3‐phosphate dehydrogenase (GAPDH) from the extreme thermophilic eubacterium Thermotoga maritima has been determined by classical automated sequence analysis of peptides derived by chemical fragmentation with cyanogen bromide and enzymatic cleavages with specific proteases.
The protein contains 332 amino acids per subunit. Its sequence is as follows: A R V A I N G F G R10 I G R L V Y R I I Y20 E R K N P D I E V V30 A I N D L T D T K T40 L A H L L K Y D S V50 H K K F P G K V E Y60 T E N S L I V D G K70 E I K V F A E P D P80 S K L P W K D L G V90 D F V I E S T G V F100 R N R E K A E L H L110 Q A G A K K V I I T120 A P A K G E D I T V130 V I G C N E D Q L K140 P E H T I I S C A S150 C T T N S I A P I V160 K V L H E K F G I V170 S G M L T T V H S Y180 T N D Q R V L D L P190 H K D L R R A R A A200 A V N I I P T T T G210 A A K A V A L V V P220 E V K G K L D G M A230 I R V P T P D G S I240 T D L T V L V E K E250 T T V E E V V A V M260 K E A T E G R K G270 I I G Y N D E P I V280 S S D I I G T T F S290 G I F D A T I T N V300 I G G K L V K V A S310 W Y D N E Y G Y S N320 R V V D T L E L L L330 K M.
Comparing the given sequence with those of the enzymes from the moderate and extreme thermophilic bacteria Bacillus stearothermophilus and Thermus aquaticus, 63% and 59% identity are observed. Alignment of the sequences of GAPDHs from a variety of sources yields one deletion (one amino acid) and one insertion (two amino acids).
Thermal stability is caused by minute adjustments of the local three‐dimensional structure. Previous ‘strategies of thermal adaptation’ in terms of preferred amino‐acid exchanges are not in accordance with the present sequence data.
Subcellular localisation of dihydrolipoamide dehydrogenase and detection of lipoic acid in bloodstream forms of Trypanosoma brucei
Authors: JACKMAN Simon A. , HOUGH David W. , DANSON Michael J. , STEVENSON Kenneth J. , OPPERDOES Fred R. 
Published: March 2005

In the long‐slender bloodstream form of Trypanosoma brucei, the enzyme dihydrolipoamide dehydrogenase exists in the absence of the 2‐oxo‐acid dehydrogenase complexes of which it is normally a component, and appears to be associated with the plasma membrane of the organism [Danson, M. J., Conroy, K., McQuattie, A. & Stevenson, K. J. (1987) Biochem. J. 243, 661–665]. In the present paper, a complete subcellular fractionation of T. brucei has been carried out and, by comparison with marker enzymes, it is confirmed that the dihydrolipoamide dehydrogenase is indeed associated with the plasma membrane. In addition, we now provide evidence that the distribution of the enzyme is over the whole surface of the membrane, including the flagellar pocket region, and that the enzyme is not found in any other cellular fraction. A study of the latency of the enzyme suggests that it is located on the cytoplasmic surface of the plasma membrane.The discovery of the presumed substrate of dihydrolipoamide dehydrogenase, lipoic acid, is reported for T. brucei. Using a biological assay involving a strain of Escherichia coli that requires lipoic acid for growth, we have found that acid‐hydrolysed extracts of T. brucei contain 1.7 (± 0.2) ng of the cofactor/mg protein. The chemical nature of the lipoic acid was confirmed by gas chromatography/mass spectrometry.
Muscle‐specific response to thyroid hormone of myosin isoform transitions during rat postnatal development
Authors: d'ALBIS Anne , CHANOINE Christophe , JANMOT Chantal , MIRA Jean‐Claude , COUTEAUX René 
Published: March 2005

Transitions from embryonic and neonatal to adult‐type‐II isomyosins are known to be related to the increase in the thyroid hormone plasma concentration during postnatal development. These transitions have been shown, however, to occur at different times, depending on the muscle, suggesting that each muscle responds differently to the thyroid hormone. We have investigated quantitatively the effects of experimental hypothyroidism and hyperthyroidism on isomyosin transitions from birth until the 45th postnatal day in eight rat muscles: diaphragm, intercostals, gastrocnemius medialis, soleus, plantar muscles of the foot, tongue muscle, levator ani and bulbocavernosus complex, and masseter.Hypothyroidism delayed the isomyosin transitions in all the muscles examined, particularly in the sexually dimorphic muscles (levator ani and bulbocavernosus complex and masseter). However, it did not eventually inhibit isomyosin transitions, indicating that the thyroid hormone was not an absolute requirement for these to occur.Hyperthyroidism had only a slight effect on isomyosin transition in the diaphragm, and accelerated such transitions in the other muscles. The transition curves of all the muscles investigated, except those of the sexually dimorphic muscles, became similar to that of the diaphragm, demonstrating that the various muscles did not display the same sensitivity to the thyroid hormone but were regulated by it in the same way. The isomyosin transitions in the sexually dimorphic muscles remained late in comparison to that in the diaphragm, which suggests a more complex regulation. The effect of hyperthyroidism was not permanent and could be reversed, by interruption of the treatment, to a greater or lesser extent depending on the muscle.In all muscles containing slow‐type‐I isomyosin, hypothyroidism had no effect on this isomyosin synthesis, whereas hyperthyroidism inhibited it. This inhibition ceased rapidly after the interruption of the treatment.
A 31P‐NMR study of bovine adrenocortical mitochondrial metabolic activities
Authors: PERRIN Anne , DEFAYE Geneviève , GUIDICELLI Claudette , CHAMBAZ Edmond , ALBRAND Jean‐Pierre , ROBY Claude 
Published: March 2005

High‐field 31P‐NMR spectroscopy has been used to study the metabolic activities of coupled bovine adrenocortical mitochondria in vitro. These differentiated organelles use oxygen as a substrate to support both oxidative phosphorylation and specific steroid hydroxylation reactions.The NMR technique allowed the resolution of two inorganic phosphate signals, attributed to the matrix and external medium phosphate pools, at low and high field, respectively. These signals were used to calculate the respective Pi concentrations and to obtain the pH of the two corresponding compartments. In addition, the NMR spectra displayed resonance signals corresponding to ADP added to the medium and to ATP synthesized during oxidative phosphorylation. NMR analysis of the mitochondrial perchloric acid extracts identified the major phosphate‐containing metabolites, namely NADP+, NAD+, phosphocholine, phosphoethanolamine, sn‐glycero‐(3)phosphocholine, AMP, ADP, ATP and Pi.Upon addition of ADP and malate to the oxygenated suspension, the kinetics of mitochondrial external Pi consumption and of ATP synthesis, along with the intra‐ and extraorganelle pH variations could be monitored over time periods of approximately 30 min, in the absence and presence of different steroid hydroxylation substrates. A major observation was that oxidative phosphorylation, which takes place in the absence of steroid, was markedly inhibited as soon as steroid hydroxylation was operating. These observations show the potential of 31P‐NMR spectroscopy in the study of metabolic activities of isolated intact mitochondrial organelles. Such an approach appears promising for further determination of the underlying mechanisms in the balance between vital oxidative phosphorylation and differentiated steroid hydroxylation which are under hormonal control in adrenocortical mitochondria as well as in other steroidogenic cell systems.
Primary structure of rat hepatocyte growth factor and induction of its mRNA during liver regeneration following hepatic injury
Authors: OKAJIMA Ai , MIYAZAWA Keiji , KITAMURA Naomi 
Published: March 2005

Overlapping cDNA clones for rat hepatocyte growth factor (rHGF) were isolated by cross‐hybridization with the cloned cDNA for human hepatocyte growth factor (hHGF) and the nucleotide sequence of the cDNA was determined. The entire primary structure of rHGF was deduced from the sequence. Comparison of the amino acid sequences between rat and human HGFs revealed that the two sequences are highly conserved throughout the protein structures, suggesting that rat and human HGFs may be functionally similar.Responses of the rHGF mRNA during liver regeneration in rats were examined by Northern blot hybridization analysis with the aid of the cDNA probe for rHGF. The mRNA levels increased in the liver and spleen but not in the kidney after administration of carbon tetrachloride. At the maximum level of induction, the rHGF mRNA increased in the liver about 4.5‐fold over its normal level. The mRNA levels also increased in the liver and spleen after administration of d‐galactosamine. On the other hand, no obvious increase of the mRNA was observed in the liver and spleen after partial hepatectomy. These observations suggest that HGF may function as a regulator of liver regeneration following hepatic injury caused by hepatotoxins.
Analysis of the binding of glyceraldehyde‐3‐phosphate dehydrogenase to microtubules, the mechanism of bundle formation and the linkage effect
Authors: SOMERS Marijke , ENGELBORGHS Yves , BAERT Johan 
Published: March 2005

The binding of glyceraldehyde‐3‐phosphate dehydrogenase (GAPDH) to microtubules was analysed by the determination of the concentration of the free enzyme in equilibrium with the complex. At low ionic strength (0.03 M) the binding data are best described by a sum of high (Kd= 0.28 μM) and low affinity (Kd= 7.14 μM) sites, showing positive cooperativity. Addition of 1 mM adenosine 5′‐[β,γ‐imido]triphosphate (p[NH]ppA) or increasing the ionic strength to 0.1 M reduces the binding constant of the high‐affinity sites considerably. Adding microtubule‐associated proteins (at I= 0.1 M) does not appreciably influence the affinities. Total stoichiometries vary over 2.1 – 1.2 tubulin dimers involved in a binding site for GAPDH.Bundling is reduced concomitantly with the reduction of the affinities and the increase of the stoichiometry to close to 1 mol GAPDH/mol tubulin dimer. The critical concentration of tubulin is practically not influenced by the binding of the enzyme. This behaviour is discussed in terms of the concept of linked functions.p[NH]ppA dissociates the bundles very rapidly. Analytical sedimentation studies showed that the dissociation of the bundles by p[NH]ppA is not due to the dissociation of the tetrameric enzyme. Bundling slows down association and dissociation of microtubules. The rate of bundle formation, after addition of GAPDH to preformed microtubules, is not dependent on the GAPDH concentration.
Carbohydrate epitope structural elucidation by 1H‐NMR spectroscopy of a new Mycobacterium kansasii phenolic glycolipid antigen
Authors: GILLERON Martine , VENISSE Anne , RIVIÈRE Michel , SERVIN Philippe , PUZO Germain 
Published: March 2005

The complete primary structure of the carbohydrate moiety of a new phenolic glycolipid antigen namely PheGl K‐IV from Mycobacterium kansasii was successfully established from only one‐ and two‐dimensional 1H‐NMR data. Among the scalar two‐dimensional techniques, correlated spectroscopy with a 45° mixing pulse and phase‐sensitive double‐quantum‐filtered correlated spectroscopy were selected, combined with two‐dimensional dipolar techniques (nuclear Overhauser effect). These techniques using milligram of quantities native PheGl K‐IV allowed the following monoacetylated tetrasaccharide to be proposed for its carbohydrate part:

image
The PheGl K‐IV shares, with the other phenolic glycolipids isolated from M. kansasii (K‐I, K‐II), a common core assigned to the lipid aglycone glycosylated by the monoacetylated trisaccharide part. It differs in the structure of the distal monosaccharide residue.
Blockage of a pump‐related calcium‐efflux pathway in light sarcoplasmic reticulum vesicles by Mops
Authors: STEPHAN Sibylle , MIGALA Andrea , HASSELBACH Wilhelm 
Published: March 2005

Mops, used as a proton buffer, specifically enhances the accumulation of calcium or strontium by light sarcoplasmic reticulum vesicles driven by ATP or dinitrophenylphosphate as energy‐yielding substrates when calcium‐precipitating agents are absent. The enhancement of ion uptake by Mops is much greater for strontium than for calcium and is further increased when potassium is replaced by sodium as the dominant monovalent cation. Mops affects neither the activity of the calcium‐ or strontium‐activated transport enzyme nor the active accumulation of calcium in the presence of oxalate, i.e. when the pump runs unidirectionally forward. Passive calcium and strontium efflux rates of approximately 40–50 nmol · mg−1· min−1 are considerably reduced when histidine/glycerophosphate or Tris/maleate are exchanged for Mops. The observed passive efflux rates and their modulation by Mops are too small, in relation to the rate of ion influx, to account for either the relatively small calcium and strontium load in the absence of precipitating agents or for its modulation by Mops.The results imply that the pump itself mediates ion efflux dependent on pump activity and the different degree of saturation of lumenal ion‐binding sites by calcium and strontium, as well as their susceptibility to Mops.
Partial release of AcPhe‐Phe‐tRNA from ribosomes during poly(U)‐dependent poly(Phe) synthesis and the effects of chloramphenicol
Authors: RHEINBERGER Hans‐Jörg , NIERHAUS Knud H. 
Published: March 2005

Poly(U)‐programmed 70S ribosomes can be shown to be 80% to 100% active in binding the peptidyl‐tRNA analogue AcPhe‐tRNA to their A or P sites, respectively. Despite this fact, only a fraction of such ribosomes primed with AcPhe‐tRNA participate in poly(U)‐directed poly(Phe) synthesis (up to 65%) at 14 mM Mg2+ and 160 mM NH+4. Here it is demonstrated that the apparently ‘inactive’ ribosomes (≧35%) are able to participate in peptide‐bond formation, but lose their nascent peptidyl‐tRNA at the stage of Ac(Phe)n‐tRNA, with n≥2. The relative loss of early peptidyl‐tRNAs is largely independent of the degree of initial saturation with AcPhe‐tRNA and is observed in a poly(A) system as well. This observation resolves a current controversy concerning the active fraction of ribosomes. The loss of Ac(Phe)n‐tRNA is reduced but still significant if more physiological conditions for Ac(Phe)n synthesis are applied (3 mM Mg2+, 150 mM NH+4, 2 mM spermidine, 0.05mM spermine).Chloramphenicol (0.1 mM) blocks the puromycin reaction with AcPhe‐tRNA as expected but, surprisingly, does not affect the puromycin reaction with Ac(Phe)2‐tRNA nor peptide bond formation between AcPhe‐tRNA and Phe‐tRNA. The drug facilitates the release of Ac(Phe)2–4‐tRNA from ribosomes at 14 mM Mg2+ while it hardly affects the overall synthesis of poly(Phe) or poly(Lys).
Regulation of Ki‐ras expression in Reuber H35 cells
Authors: CHAN Sai‐On , WONG Susanna Siu‐Chun , YEUNG Desmond Chak‐Yew 
Published: March 2005

Glucagon at a low concentration has a stimulatory effect on Ki‐ras expression, whereas, at high concentrations, the hormone suppresses the level of the Ki‐ras transcripts. Incubation of the hepatoma cells with 10 μM dibutyryl cyclic AMP results in suppression of Ki‐ras expression but the phorbol ester, 21‐O‐tetradecanoylphorbol 13‐acetate (TPA) causes an increase. Down regulation of protein kinase C by prolonged exposure of hepatoma cells to TPA causes a dramatic decrease in the glucagon‐stimulated effect on Ki‐ras expression. The presence of diacylglycerol for 2 h in the culture medium results in a significant increase in Ki‐ras expression, while treatment of the cells with 1‐(5‐isoquinolinylsulphonyl)‐2‐methylpiperazine, a potent inhibitor of protein kinase C, leads to a dramatic reduction. The calcium ionophore, A23187 is able to stimulate Ki‐ras expression, whereas, addition of verapamil or EGTA results in its suppression.The present findings suggest that the inductive effect of glucagon on Ki‐ras expression at low concentrations is via the activation of protein kinase C which causes phosphorylation of some regulatory proteins that may eventually affect the level of Ki‐ras mRNA. The suppressive effect of glucagon at higher concentrations is via an increase in cAMP through activation of adenylate cyclase.