bluuebunny/crossref_metadata_2025 huggingface dataset:
Acid endopeptidase activity of human myelin, elicited by using exogenous myelin basic protein as enzyme substrate
Authors: Berlet Hans H.
Published: July 2002

Purified human myelin was incubated with exogenous myelin basic protein (MBP) at pH 4.0 to see if there is acid proteinase activity associated with myelin. Following incubation for 12 h up to 70% of MBP was degraded. On electrophoresis peptide fragments of MBP between 15.8 and 9.4 kDa were consistent with an endopeptic cleavage of MBP. Unlike the exogenous substrate MBP associated with myelin was only slightly degraded under the experimental conditions used. The results show that proteinase activity associated with isolated myelin may be elicited and further evaluated by using MBP as enzyme substrate.
Improved purification and N‐terminal sequence of the 33‐kDa protein in spinach PS II
Authors: Yamamoto Yasusi , Hermodson Mark A. , Krogmann David W.
Published: July 2002

Significant improvement was attained in the purification of a peripheral 33‐kDa protein from spinach chloroplasts by the use of butanol/water phase partitioning. Over 30% of the 33‐kDa protein in the chloroplasts was isolated in a short time. The amino acid sequence for the first 37 residues from the N‐terminus was determined by the automated sequencing of the purified protein.
Fine tuning of ribosomal accuracy
Authors: Ninio Jacques
Published: July 2002

If the rate constant for peptide bond formation were high just after an amino acid incorporation and occasionally switched to a lower value afterwards, then the ribosome could compensate for tRNA imbalance specifically at hungry codons. A rigorous analysis of the scheme proves its effectiveness. For instance, a 10‐fold reduction in cognate tRNA concentration may increase the error rate by only a factor of two.
Characterization and quantitative topographical distribution of salmon calcitonin‐binding sites in rat kidney sections
Authors: Bouizar Z. , Rostène W.H. , Moukhtar M.S. , Milhaud G.
Published: July 2002

Renal binding sites for labelled salmon calcitonin (sCT) were studied using cryostat sections and autoradiography. Increasing concentrations of unlabelled sCT inhibited 125I‐sCT binding. 125I‐sCT bound to a single site with a K
d of 2 nM and a number of sites of 220 protein. Mammalian calcitonins had low affinities and peptides unrelated to CT were devoid of any significant affinity for 125I‐sCT receptors. Auto‐radiograms disclosed a high concentration of 125I‐sCT receptors mainly located in the outer medulla and heterogeneously in the renal cortex. The distribution of specific binding sites is in agreement with the current concepts of renal action of calcitonin.
Cytoplasmic pH and glycolysis in the Dictyostelium discoideum cell cycle
Authors: Aerts R.J. , Durston A.J. , Moolenaar W.H.
Published: July 2002

Lactate production measurements during the cell cycle of synchronized populations of Dictyostelium discoideum cells reveal cyclic variations in glycolysis which correspond with pHi oscillations which were discovered by us previously [(1985) Cell, in press]. Aerobic lactate production varies about 6‐fold during the cell cycle and the lactate maxima correlate with (~ 0.25 pH unit) cyclic increases in pH. However, artificially altering pHi using weak acids or bases does not influence the rate of lactate production in asynchronous cell populations. This result suggests that the cyclic variations in pHi and those in glycolytic rate are not causally related events.
Bradykinin stimulates GTP hydrolysis in NG108‐15 membranes by a high‐affinity, pertussis toxin‐insensitive GTPase
Authors: Grandt Rüdiger , Greiner Claudia , Zubin Palmira , Jakobs Karl H.
Published: July 2002

In membranes of neuroblastoma × glioma hybrid (NG108‐15) cells, bradykinin (EC50 ≅ 5 nM) stimulates GTP hydrolysis by a high‐affinity GTPase (Km
≅ 0.2 μM). The octapeptide, des‐Arg9‐bradykinin, was inactive. Stimulation of GTP hydrolysis by bradykinin and an opioid agonist was partially additive. Treatment of NG108‐15 cells with pertussis toxin, which inactivates Ni, eliminated GTPase stimulation by the opioid agonist but not by bradykinin. The data suggest that bradykinin activates in NG108‐15 membranes a guanine nucleotide‐binding protein which is not sensitive to pertussis toxin and which may be involved in brady‐kinin‐induced stimulation of phosphoinositide metabolism in these cells.
Structure of protein‐deficient 50 S ribosomal subunits
Authors: Gongadze G.M. , Selivanova O.M. , Gudkov A.T. , Vasiliev V.D.
Published: July 2002

The complex of 23 S ribosomal RNA with the nine core proteins L2, L3, L4, L13, L17, L20, L21, L22 and L23 obtained either by the disassembly procedure or by reconstitution has been studied by electron microscopy. This complex is found to be very similar to the intact 50 S subunit both in size and in shape.
Inactivation of cytosolic aspartate aminotransferase accompanying modification of Trp 48 by N‐bromosuccinimide
Authors: Nagashima Fujio , Tanase Sumio , Morino Yoshimasa
Published: July 2002

Reaction of N‐bromosuccinimide with pig heart cytosolic aspartate aminotransferase led to loss of the enzymatic activity. Chemical analysis indicated the modification of two tryptophan residues. At a low ratio of N‐bromosuccinimide to enzyme, oxidation of Trp 122 occurred without affecting the enzymatic activity. Increase in the ratio resulted in the oxidation of Trp 48 with a concomitant decrease in enzyme activity. The modified enzyme did not react with substrates and their analogs. Trp 48 is not within the active site but in the hinge region linking the large domain of the enzyme to the small domain that shows dynamic movement upon binding substrates. The present result suggests that oxidation of Trp 48 may impair the structural integrity of the interdomain interface.
Identification of a 27‐kDa enkephalin‐containing protein associated with bovine adrenal medullary chromaffin granule membranes by immunoblotting
Authors: Birch Nigel P. , Davies Anne D. , Christie David L.
Published: July 2002

An antiserum which recognizes high molecular mass enkephalin‐containing proteins was used to compare proenkephalin intermediates in both the soluble and membrane components of bovine adrenal chromaffin granules by immunoblotting. While a range of molecular mass forms were identified in the soluble lysate the major form in the membranes corresponded to a 27‐kDa enkephalin‐containing protein. Enzymic digestion of bands of 27‐kDa material and quantitation of the enkephalin released showed that 22% of this material was membrane‐associated. High concentrations of chaotropic agents were required to extract this material from the membranes. Association of hormone and neuropeptide precursors with membrane components may by important for targeting of precursors to secretory granules or correct processing.
Polymerization of β‐like actin from scallop adductor muscle
Authors: Khaitlina S.Yu.
Published: July 2002

Scallop adductor muscle β‐like isoactin differs from rabbit skeletal muscle α‐actin in the rate, extent and critical concentration of polymerization. The difference is temperature‐ and [KCl]‐dependent. In the presence of DNase I scallop actin was shown to be depolymerized more rapidly than rabbit actin. It was suggested that the polymers formed by β‐actin are less stable than those formed by α‐actin.
A hybrid protein between IFN‐γ and IL‐2
Authors: Seno Masaharu , Hinuma Shuji , Onda Haruo , Igarashi Koichi
Published: July 2002

The complementary DNAs encoding human interferon‐γ (IFN‐γ) and human interleukin‐2 (IL‐2), two different proteins involved in the same immune system, were fused to code a hybrid protein, which was expressed in E. coli to investigate the interactions of these two proteins at the molecular level. Through immunoprecipitation analysis, this protein was revealed to be of about 31 kDa, which was expected from nucleotide sequencing, and to have the antigenicities of both IFN‐γ and IL‐2. The extract from bacteria expressing this hybrid protein showed at least two biological activities: an antiviral activity derived from IFN‐γ and the ability to support the growth of natural killer (NK) cells derived from IL‐2. Comparing the enhancement of NK cell activity of this hybrid protein with IFN‐γ and IL‐2, this hybrid protein appears to conserve each activity almost completely without diminishing the other.
Interaction of a macromolecular polyanion, dextran sulfate, with human hemoglobin
Authors: Sacco Daniel , Dellacherie Edith
Published: July 2002

Interactions of dextran sulfate with amino groups of oxy‐ and deoxyhemoglobin were followed by both potentiometric measurements between pH 6 and 7.3 and oxygen‐binding studies. The uptake of protons observed upon addition of dextran sulfate to hemoglobin shows that the interaction with the deoxy form is strong and that the main site is probably located in the phosphate‐binding β‐cavity, whereas the interaction with the oxy form is more diffuse, probably with a great number of relatively weak binding sites. The influence of dextran sulfate on the oxygen dissociation curve of hemoglobin confirms these findings, as the effect of the polymer is to lower hemoglobin affinity for oxygen to a great extent, which proves that it stabilizes the deoxy form more strongly than the oxy one.
Protein phosphorylation in isolated mitochondria and the effects of protein kinase C
Authors: Backer J.M. , Arcoleo J.P. , Weinstein I.B.
Published: July 2002

When isolated intact rat liver mitochondria are incubated with [γ‐32P]ATP the major phosphorylated proteins are those of 47 and 36 kDa. Phosphorylation of the 47 kDa protein, but not of the 36 kDa protein, is inhibited by carboxyatractyloside, an inhibitor of mitochondrial ATP uptake, while phosphorylation of the 36 kDa protein is inhibited by various uncouplers and an inhibitor of mitochondrial respiration. Addition of purified protein kinase C to the isolated mitochondria leads to the phosphorylation of 69, 37 and 17 kDa proteins. As with other substrates for protein kinase C, phosphorylation of these proteins is dependent on Ca2+ and markedly stimulated by various tumor promoters.
The nucleotide sequences of chloroplast 4.5 S rRNAs from four species of plants, celery (Apium graveoleus), barley (Hordeum vulgare), Chinese chive (Allium tuberosum) and dayflower (Commelina communis)
Authors: Zhen-Qi Cheng , Hong Zhang , Guo-Ya Li , Xiao-Yang Liu
Published: July 2002

Four chloroplast 4.5 S rRNAs were isolated from the respective plant leaves by a simple method. The complete nucleotide sequences were determined using rapid gel sequencing techniques. The sequences are highly conserved among chloroplast 4.5 S rRNAs. The 4.5 S rRNAs of celery, Chinese chive and dayflower are 103 nucleotides long and that of barley 95 nucleotides long. The 7‐nucleotide sequence from position 28–34 is absent in monocotyledon barley 4.5 S rRNA but is present in monocotyledon Chinese chive and day‐flower.
Specific lysis of noradrenergic synaptosomes by an antiserum to dopamine‐β‐hydroxylase
Authors: Docherty Maureen , Bradford H.F. , Joh T.H.
Published: July 2002

An antiserum to dopamine‐β‐hydroxylase purified from bovine adrenal medulla, acting in the presence of complement, caused the release of 12% of lactate dehydrogenase, 20% of tyrosine hydroxylase activity, and 40% of noradrenaline (NA) content from synaptosomes prepared from rat brain cerebral cortex. Uptake of [3H]NA was reduced by 54%. Anti‐serum alone or complement alone were without action. The antiserum plus complement had no effect on choline uptake and did not release choline acetyltransferase, glutamate decarboxylase, dopamine or 5‐hydroxytryptamine. These results suggest selective lysis of noradrenergic terminals had occurred. An enhancement of lysis was not observed when synaptosomes were stimulated with 75 K+ and exposed to a sub‐maximal dose of antiserum, plus complement.
Two‐dimensional lattices of porin diffract to 6 Å resolution
Authors: Büldt Georg , Mischel Maja , Hentschel Manfred P. , Regenass Martin , Rosenbusch Jürg P.
Published: July 2002

Porin (the product of gene ompF) is an integral membrane protein (M
r 36 500) of the outer membrane of Escherichia coli (strain BE). The protein has been purified to homogeneity and reconstituted in dimyristoyl‐lecithin. Oriented specimen on a flat surface yielded X‐ray diffraction pattern, originating from the two‐dimensional protein lattice, to a resolution reaching 6 Å. Although these powder rings are broad compared to corresponding diffraction patterns from purple membranes of Halobacterium halobium, porin is the first reconstituted integral membrane protein which shows diffraction to this resolution.
Novel specificity of an endoribonuclease of yeast
Authors: Stevens Audrey
Published: July 2002

Investigations of the cleavage specificity of a cyclizing endoribonuclease of yeast show that it hydrolyzes Y‐A bonds of single‐stranded regions of RNA and that oligonucleotides are hydrolyzed poorly. The stringent specificity of the enzyme suggests that it may be useful for generating specific fragments of polyribonucleotides. An isolation technique employing ethanol extraction has now been used to isolate the enzyme free of other RNases, and the studies show that it is a small protein, 10 kDa or less in molecular mass.
Inhibition of the mitochondrial tricarboxylate carrier by arginine‐specific reagents
Authors: Stipani I. , Zara V. , Zaki L. , Prezioso G. , Palmieri F.
Published: July 2002

The effect of arginine‐specific reagents on the activity of the partially purified and reconstituted tricarboxylate carrier of the inner mitochondrial membrane has been studied. It has been found that 1,2‐cyclohexanedione, 2,3‐butanedione, phenylglyoxal and phenylglyoxal derivatives inhibit the reconstituted citrate/citrate exchange activity. The inhibitory potency of the phenylglyoxal derivatives increases with increasing hydrophilic character of the molecule. Citrate protects the tricarboxylate carrier against inactivation caused by the arginine‐specific reagents. Other tricarboxylates, which are not substrates of the carrier, have no protective effect. The results indicate that at least one essential arginine residue is located at the substrate‐binding site of the tricarboxylate carrier and that the vicinity of the essential arginine(s) has a hydrophilic character.
Aromatic ring cleavage of β‐O‐4 lignin model dimers without prior demeth(ox)ylation by lignin peroxidase
Authors: Umezawa Toshiaki , Higuchi Takayoshi
Published: July 2002

Methyl oxalate of arylglycerol was formed as an aromatic ring cleavage product in degradation of arylglycerol‐β‐aryl ether (β‐O‐4) type lignin substructure model dimers by extracellular lignin peroxidase of Phanerochaete chrysosporium. The enzymatic cleavage of arylglycerol‐β‐(o‐[2H3]methoxyphenyl) ether indicated that the methyl group of the methyl ester was derived from the methoxy group of the β‐O‐4 model dimer. It is thus concluded that demeth(ox)ylation was not essential for the enzymatic aromatic ring cleavage of the methoxylated aromatic substrates, β‐O‐4 lignin substructure models.
Characterization of the distribution of potential short restriction fragments in nucleic acid sequence databases
Authors: Tung Chang-Shung , Burks Christian
Published: July 2002

We have searched the GenBank nucleic acid sequence database for potential short restriction fragments. All possible oligonucleotides up to length five are found at least once flanked by known restriction recognition patterns. Thus, searches in the database for a specific sequence corresponding to a desired oligonucleotide would often point to one or more sources of short, retrievable fragments containing that sequence. These results underscore the potential of nucleic acid sequence databases in planning experiments.
Regulation of ATP hydrolase activity of the F0‐F1 complex of rat‐liver mitochondria during early hepatic regeneration
Authors: Buckle Malcolm
Published: July 2002

Submitochondrial particles prepared from rat liver in the early phase of hepatic regeneration possess a reduced F1 content with respect to F0 in intact F0F1‐H+‐ATPase complexes. Analysis of ATP hydrolysis showed a significant difference in both ESMP and isolated F1 with regard to the higher affinity K
m values (K
m,1) obtained from Eadie‐Hofstee plots. Both ESMP and F1 from regenerating rat liver showed much lower apparent K
m,1 values (0.04 and 0.03 mM, respectively) than the corresponding controls (0.08 mM for both ESMP and F1). Data presented here show that the residual F1 moieties have an altered kinetic pattern with regard to the competitive inhibitor adenosine 5'‐[β,γ‐imido]triphospate (K
1 ESMP from regenerating rat liver = 0.67 μM, K
1 ESMP from control rat liver = 2.03 μM). This difference in affinity for [β,γ‐imido]‐ATP is also seen in isolated F1(K1 regenerating rat liver = 0.04 μM, K
1 control rat liver = 0.22 μM). These data indicate that during the disruptive retrodifferential phase of hepatic regeneration, changes at the level of surviving F1 sectors of the F0‐F1 ATPase may play a physiological role in preventing ATP hydrolysis in vivo in the brief period of low ΔμH+, induced by the presence of non‐F1‐associated F0 proton‐conducting pathways.
Fructose 2,6‐bisphosphate and glucose 1,6‐bisphosphate in erythrocytes during chicken development
Authors: Espinet Carme , Bartrons Ramon , Carreras José
Published: July 2002

In contrast to mammalian erythrocytes, chicken erythrocytes contain fructose 2,6‐bisphosphate at levels (0.5 cells) similar to those of 2,3‐bisphosphoglycerate (1.2 nmol/109 cells) and slightly lower than those of glucose 1,6‐bisphosphate (5.2 cells). In chick embryo erythrocytes the levels of both fructose 2,6‐bisphosphate and glucose 1,6‐bisphosphate are much lower. They begin to increase at hatching and reach the levels in chicken in a few days.
Origins of stereospecificity in DNA damage by anti‐benzo[a]pyrene diol‐epoxides
Authors: Pearl Laurence H. , Neidle Stephen
Published: July 2002

A general computational procedure for the modelling of intercalated DNA‐ligand complexes has been developed, and is used here to model intercalated complexes of the (+)‐anti and (‐)‐anti enantiomers of benzo[a]pyrene diol‐epoxide (BPDE) with cytosine‐3',5'‐guanosine double‐stranded DNA sequences (dCpG). Results are presented indicating differences between the behaviours of the two enantiomers which have implications for the understanding of the stereospecificity of DNA strand breakage by benzo[a]pyrene diol‐epoxides.
Coat protein blocks the in vitro transcription of the virion RNAs of alfalfa mosaic virus
Authors: Houwing Corrie J. , Jaspars E.M.J.
Published: July 2002

Using a method described by Miller and Hall [(1983) Virology 125, 236‐241] for brome mosaic virus, we succeeded in obtaining preparations of an alfalfa mosaic virus specified RNA polymerase that is totally dependent on added RNA. The enzyme makes full‐size transcripts on each of the four virion RNAs. Transcription is inhibited by small amounts of coat protein subunits. This is in accordance with a model proposed by Nassuth and Bol [(1983) Virology 124, 75‐85] which says that in a late stage of the infection cycle the coat protein is a factor that regulates the minus‐strand RNA synthesis in a negative sense.
Receptor binding, cGMP stimulation and receptor desensitization by atrial natriuretic peptides in cultured A 10 vascular smooth muscle cells
Authors: Neuser Dieter , Bellemann Peter
Published: July 2002

Receptor characteristics for atrial natriuretic peptides (ANP) were demonstrated in the permanent tissue culture system of vascular smooth muscle cell (VSMC) line. 125I‐ANP exhibited reversible and saturable binding to A10 rat VSMC, and the equilibrium dissociation constant, K
d was 157 pM and maximal binding capacity, .B
max amounted to 115 of protein. Binding of the 125I‐ligand was highly specific for certain potently displacing ANP analogues with inhibition constants (K
i values) in the nano‐ or even subnanomolar concentration range. Pretreatment of VSMC with ANP yielded receptor desensitization to one half of the ANP receptor density, and supports the conclusion of receptor autoregulation by ANP in A10 VSMC. The receptor coupled intracellular cGMP system was stimulated, and thus demonstrates the application of A10 cells as a model for the study of ANP receptor interaction involved in vascular smooth muscle relaxation.
Is phospholipid a required cofactor for the activity of mammalian signal peptidase?
Authors: Uchida Masahide , Watanabe Yasuhiro , Fujimoto Yukio
Published: July 2002

To determine whether phospholipid is required for the activity of mammalian signal peptidase, the enzyme was partially purified from porcine pancreas and then extensively freed of phospholipid by SP‐Sephadex C‐50 chromatography. The delipidated enzyme showed signal peptidase activity, with a low concentration of detergent. Phospholipid was found to release the enzyme from the inhibition due to excess detergent.
Norepinephrine with its precursors and their antagonists haloperidol and phentolamine interact with dye free radicals in opposite ways
Authors: Marinov B.S.
Published: July 2002

In the reactions with dye free radicals, catecholamines exhibited reversible electron donor and acceptor properties with the effectiveness increasing from tyrosine to norepinephrine. The physiological antagonists haloperidol and phentolamine showed opposite patterns of behaviour in the same reactions, changing their properties as acceptors to electron donors. The regularity observed is similar to that demonstrated earlier by a variety of Na+ and Ca2+ channel modulators.
An osmotic model for the fusion of biological membranes
Authors: Lucy Jack A. , Ahkong Quet F.
Published: July 2002

A molecular model for fusion‐fission reactions in membranes is proposed that is based on data from studies on artificially induced cell fusion and on the behaviour of phospholipid bilayers: it is put forward as a framework for further investigations into this fundamental property of biological systems.
Existence of antizyme and ornithine decarboxylase‐antizyme complex in RK13 kidney cells
Authors: Murakami Yasuko , Marumo Miya , Hayashi Shin-ichi
Published: July 2002

It has been reported that ‘antizyme’, a protein inhibitor of ornithine decarboxylase (ODC) induced by its product, is not found in rat or mouse kidney. We determined whether antizyme was present in rabbit kidney cells (RK13) in culture. Antizyme could be induced in these cells by putrescine treatment, a substantial portion being in the particulate fraction in contrast with hepatic antizyme. Furthermore, ODC‐antizyme complex was present even in untreated cells. Pretreatment of cells with putrescine increased the relative amount of ODC‐antizyme complex and accelerated decay of ODC. These results support the ubiquitous existence of antizyme and its role in ODC degradation.
Deficiency of neutrophil membrane antigen detected by monoclonal antibody in rheumatoid arthritis
Authors: Traniello Serena , Spisani Susanna , Gavioli Riccardo , Dovigo Livio , Baricordi Roberto O. , Sensi Alberto , Damiani Guido
Published: July 2002

Monoclonal antibodies (mAbs) against cell surface antigens and receptors are instrumental in defining specific membrane markers. mAbs GF26.7.3 and MF25.1 against human neutrophils modulated the activation mechanism of superoxide anion production induced by formyl‐peptide and PMA in all subjects. However, treatment with mAb MF 25.1 binds is absent in rheumatoid conditions. This confirms our previous data showing that defective expression of membrane components is associated with neutrophil dysfunction.
Purification of an enzyme aggregate containing 3′, 5′‐cyclic‐nucleotide phosphodiesterase and nucleotidase
Authors: Böldicke Thomas W. , Schubert-Rehberg kristiane , Wombacher Helmut
Published: July 2002

Several steps of purification (octyl‐Sepharose chromatography,Blue Sepharose 6B Chromatography and sucrose density gradient centrifugation) led to a highly purified aggregate of the enzymes, 3′, 5′‐cyclic‐nucleotide phosphodiesterase (PDE) and nucleotidase. The purfied enzyme aggregate showed an S value of 7.3 (SE±0.3, n = 10). Further analysis by SDS‐polyacrylamide gel electrophoresis (PAGE) reavealed two proteins near 67 and 60 kDa. Dissocation of the 7.3 S enzyme aggregate showed a 3.6 S PDE form and a nucleotidase form at 4.2 S. Additionally, higher S value forms of the necleotidase up to 17 S have been observed. Apparently, they had formed by self‐association. SDS‐PAGE of the 17 S nucleotidase form showed only one hand at 67 kDa. This was taken as evidence for the homogenity of the 17 S nucleotidase form and the self‐association of the nuleotidase after dissociation from the 7.3 S enzyme aggregate. Furthermore, from this it could be concluded that the 67 kDa protin of the 7.3 S enzyme aggregate should be identified with the nucleotidase, and thus the 60 kDa band represents the PDE.
Isolation and partial characterization of 2‐μm yeast plasmid as a transcriptionally active minichromosome
Authors: Silva Jesus , Zinker Samuel , Gariglio Patricio
Published: July 2002

Yeast cell extracts from 2‐μm‐containing strains (cir+) showed higher transcriptional activity than their corresponding isogenic sets (cir°). These extracts were used to purify transcriptionally active 2‐μm minichromosomes in a sucrose gradient. Minichromosomes were transcribed in vitro and, employing hybridization techniques, the RNA synthesized was shown to present 2‐μm‐specific sequences. This model system should permit the direct study of transcriptionally active eucaryotic chromatin.
Proton magnetic resonance of the bovine spleen green heme‐protein
Authors: Ikeda-Saito Masao , Inubushi Toshiro
Published: July 2002

The ferric spleen green heme‐protein exhibits hyperfine‐shifted proton resonances between 90 and 20 ppm for the high‐spin resting form and the chloride complex, and between 46 and −9.4 ppm for the low‐spin nitrite complex. The proton NMR spectral profile of the enzyme is similar to that of lactoperoxidase, but different from those of common heme‐proteins. The appearance of a resonance at 76 ppm in the ferrous enzyme shows the presence of a proximal histidine residue linked to the iron. The proton relaxation rates of bulk water indicate that chloride binds to the sixth position of the iron in the chloride complex of the enzyme.
Decrease of cellular ATP by dihexanoylglycerol may limit responses to protein kinase C activation
Authors: Dawson J. , Thompson N.T. , Bonser R.W. , Hodson H.F. , Garland L.G.
Published: July 2002

1,2‐sn‐Dihexanoylglycerol (HHG) reduced the ATP content of HL‐60 cells. This concentration‐related (10–100 μM) effect reached a maximum of over 90%, was enantiomerically specific and not accompanied by release of lactate dehydrogenase. Oleoylacetylglycerols (3–100 μM) had no effect on ATP levels while phorbol dibutyrate (PDBu, 0.01–1 μM) decreased ATP content of HL‐60 cells by up to 40%. Responses stimulated by HHG became limited as the concentration was increased above 10 μM, this being manifest as either a low maximum response compared to PDBu (superoxide release) or a bell‐shaped concentration‐effect curve (degranulation). HHG (30–100 μM) inhibited PDBu‐stimulated superoxide release, this inhibition being enantiomerically specific. It is probable that the effect of HHG on ATP content impairs cellular reponses mediated through protein kinase C activation.
The kinetic effects of in vitro phosphorylation of rabbit muscle enolase by protein kinase C
Authors: Nettelblad Folke A. , Engström Lorentz
Published: July 2002

Rabbit muscle enolase was found to be phosphorylated in vitro by calcium‐activated phospholipid‐dependent protein kinase. The extent of incorporation (about 0.6 mol/mol subunit) and the apparent K
m for the reaction (3.0 μM subunit) were determined. Kinetic studies on the unphosphorylated and phosphorylated enzymes displayed an unexpected effect of phosphorylation resulting in activation of the forward reaction and inhibition of the backward one.
ESR signals from stimulated and resting porcine blood neutrophils
Authors: Hata-Tanaka Akiko , Chiba Tsukasa , Kakinuma Katsuko
Published: July 2002

The NADPH oxidase in neutrophils was specifically solubilized from membrane vesicles of porcine blood neutrophils and rapidly concentrated by immunoprecipitation with cross‐reacting anti‐P‐450 reductase IgG. The precipitates from both myristic acid‐stimulated and resting cells contained one third of the cytochrome b‐558 and were slightly contaminated with myeloperoxidase. The immunoprecipitate from stimulated cells gave rhombic high‐spin ESR signals of a heme at g = 6.47 and 5.49, which were insensitive to KCN, whereas the preparation from resting cells did not give these signals. The rhombic high‐spin signals are discussed in view of the participation of cytochrome b‐558 in the NADPH oxidase system.
The structure of barley stripe mosaic virus double‐stranded RNAs
Authors: Dolja Valerian V. , Atabekov Joseph G.
Published: July 2002

The terminal structures of the double‐stranded replicative forms (RFs) of barley stripe mosaic virus (BSMV) RNAs 1–3 have been investigated. All three BSMV RFs have identical right‐hand ends but unique left‐hand ends. The plus (+) strands of RFs lack the 3′‐ultimate A typical for the encapsidated BSMV RNAS. The 3′‐termini of the minus (−) strands contain an unpaired G. It was demonstrated that the internal poly(A) tract of BSMV genome has an equivalent poly(U)‐counterpart in the RF (−) strands. The possible role of these peculiarities of BSMV RF structure in RNA replication is discussed.
The ATP‐dependent generation of membrane potential by sub‐bacterial vesicles from the marine bacterium, Vibrio alginolyticus
Authors: Smirnova Irina A. , Vulfson E.N. , Kostyrko V.A.
Published: July 2002

Addition of ATP leads to the accumulation of the permeant anion PCB− by sub‐bacterial vesicles from Vibrio alginolyticus. This accumulation is caused by Δψ generation by ATPase, the effect being inhibited by CCCP, gramicidin D and DCCD. Δψ values may be increased by incubation of sub‐bacterial vesicles at room temperature and with the protein fraction isolated according to Beechey et al. [(1975) Biochem. J. 148, 533–537] from another portion of the sub‐bacterial vesicles. Δψ generation is observable only in the presence of Mg2+ at high concentrations (optimum ≈ 30 mM). Proceeding from experimental data we assume that Mg2+ reduces passive H+ conductivity of the vesicle membranes. Thus, a Δψ‐generating ATPase has been shown for the first time in V. alginolyticus membranes.
Apolipoprotein A‐I‐binding protein from human term placenta Purification and partial characterization
Authors: Keso Leila , Lukka Matti , Ehnholm Christian , Baumann Marc , Vihko Pirkko , Olkinuora Maritta
Published: July 2002

A protein that binds to the main apoprotein, apoA‐I, of human high density lipoprotein (HDL) has been isolated from human placenta. Ligand blotting after SDS gel electrophoresis indicated that the 120 kDa protein in the absence of reducing agents binds apoA‐I. If gel electrophoresis was performed under reducing conditions two main bands, approx. 50 and 30 kDa that did not bind apoA‐I, were evident. In an enzyme‐linked immunosorbent assay the binding protein specifically bound apoA‐I, delipidated or as HDL. ApoA‐II, apo E and LDL did not compete with apoA‐I for binding to this protein.
Prediction of exposure degree diagram and sites of limited proteolysis in globular proteins as an approach to computer‐aided design of protein bioregulators with prolonged action
Authors: Rodionov M.A. , Galaktionov S.G. , Akhrem A.A.
Published: July 2002

In order to prolong the lifetime of protein bioregulators in blood it is possible to engineer analogs with protected sites of limited proteolysis. To determine the sites, primarily accessible to trypsin‐like proteases, a computer procedure has been developed, including a prediction algorithm, to produce the residue diagram of a globular protein and a discriminant algorithm to determine the sites most liable to proteolysis. The accuracy of prediction of amino acid residue exposure is characterised by correlation coefficients between experimental and theoretical exposure values, the coefficients being about 0.7 as calculated for 10 globular proteins. The classification of Arg and Lys residues into two groups, susceptible or insusceptible to protease, has an error percentage of about 25.
Co‐evolution of base composition and codon usage in Xenopus laevis and human globin genes with long‐range DNA organization of their genome
Authors: Moreau J. , Scherrer K.
Published: July 2002

Eucaryotic DNA is punctuated by many A+T‐rich segments that we named A+T‐rich linkers. Two types of these A+T‐rich linkers can be distinguished: (i) isolated A+T‐rich linkers, and (ii) A+T‐rich linkers crowded in clusters. We have analysed the distribution of A+T‐rich linker across the α‐ and β‐globin gene domain in Xenopus laevis and human genomes using isodenaturation and electron microscopy. Comparison of our data with those previously obtained for the avian globin genes leads us to conclude that genes can be harboured indifferently in either domain. A correlation is established between the presence of A+T‐rich linker inside introns and flanking regions and the A+T content of the coding sequence. For the coding sequence, a high A+T content is strongly correlated with high A+T content in the codon's third position and weakly in the first position.
Vitamin K‐dependent carboxylase Possible role for thioredoxin in the reduction of vitamin K metabolites in liver
Authors: van Haarlem L.Johan M. , Soute Berry A.M. , Vermeer Cees
Published: July 2002

In the liver vitamin K epoxide, which is produced during the posttranslational carboxylation of proteinbound glutamic acid residues, is recycled by the action of one or more dithiol‐dependent reductases. In vitro synthetic dithiols may serve as a cofactor for these enzymes, but the physiological reluctant has not yet been found. In this paper we report that in vitro the commercially available thioredoxin/thioredoxin reductase from E. coli can replace the synthetic dithiols during the various reactions of the vitamin K cycle. Based on the assumption that in vivo thioredoxin also plays a role in the regeneration of vitamin K hydroquinone from the epoxide, an extension of the generally accepted vitamin K cycle is proposed.
The complete amino acid sequence of proapolipoprotein A‐I of chicken high density lipoproteins
Authors: Yang Chao-yuh , Gu Zi-Wei , Patsch Wolfgang , Weng Shi-ai , Kim Tae W. , Chan Lawrence
Published: July 2002

The complete amino acid sequence of proapolipoprotein (proapo) A‐I of chicken high density lipoproteins was determined by sequencing overlapping peptides produced by trypsin, S. aureus V8 protease, and cyanogen bromide cleavage. There are 240 amino acid residues in mature chicken apoA‐I. By direct sequence analysis of a cyanogen bromide peptide, we also determined the sequence of a 6‐amino‐acid prosegment which is present at approx. 10% the molar amount of the mature peptide in chicken plasma. Sequence comparison among apoA‐I from chicken, human, rabbit, dog and rat, and secondary structure analysis indicate that while the degree of sequence homology is only moderate ( < 50% between chicken and man), there is good conservation of apoA‐I secondary structure, especially in the N‐terminal two‐thirds of the protein in these widely separated species.
Purification and characterization of a recombinant human IgE Fcε fragment lacking the C4 domain
Authors: Ikeyama Shuichi
Published: July 2002

Complementary DNA of human lgE Icε fragment (residues 226–480) lacking the Cϕ domain was expressed in Escherichia coli and the product was purified by immunoaffinity chromatography on a monoclonal antibody (E12 0.02)‐Affi‐Gel 10 column. About 1.8 mg of an apparent dimer and 5.9 mg of a monomer were obtained from 65 g E. coli cells with 9.3% recovery. The purified products were found to lack more than half of the COOH‐terminal portion of the Cε3 domain. The apparent dimer showed high immunological specific activity (3.6 × 106 U/mg protein) comparable to that of natural human IgE when measured by commercial human IgE determination kits.
Phosphorylation by cyclic AMP‐dependent protein kinase does not affect the association of ATP citrate‐lyase with isolated mitochondria
Authors: Lin Tianwei , Bridger William A.
Published: July 2002

ATP citrate‐lyase is known to be a substrate for various protein kinases, but the functional role, if any, of kinase‐directed phosphorylation of this enzyme has not been identified. Recently, Strålfors [(1987) J. Biol. Chem. 262, 11486–11489] has suggested that effects on the association of this enzyme with mitochondria may account for the observed ability of isoproteronol or insulin to promote immobilization of ATP citrate‐lyase in permeabilized cells. Here we report studies involving phosphorylation of the pure enzyme in vitro using cyclic AMP‐dependent protein kinase. We show that phosphorylation has no significant effect on the fraction of the enzyme that may be bound to isolated mitochondria.
Statistics of the photon distribution in the set of photosynthetic antenna domains
Authors: Mineyev A.P. , Razjivin A.P.
Published: July 2002

The distribution of the excitations caused by the pump laser pulse in the domains of light‐harvesting antenna is shown to be the hypergeometrical one. The experimental curves of the photo‐oxidation of reaction centers versus the light intensity of pico‐second pulses measured for two distinct samples with monocentral antenna domains are in excellent agreement with the statistical distribution predicted.
Isolation and structural characterization of porcine coupling factor 6 from intestinal tissues
Authors: Chen Zheng-wang , Mutt Viktor , Barros-Söderling Jane , Jörnvall Hans
Published: July 2002

A polypeptide purified from an extract of thermostable, porcine intestinal peptides was found to correspond to coupling factor 6, previously known as a component of the mitochondrial oxidative phosphorylation system. The intestinal presence of this peptide offers a new source for preparation of the component in large quantities, and possibly suggests further functions of the polypeptide. Amino acid sequence analysis of this porcine form reveals it to be identical to the bovine form, except for two replacements, at position 62 (Thr in the porcine, Phe/Thr in the bovine form), and position 70 (Ala/Val). The extensive conservation suggests strict structural constraints on the functional properties of the polypeptide.
Effects of protein‐free diet in amino acid homeostatis of rat blood plasma and gut contents
Authors: Suzić Slavica , Radunović Ljubinka , Janković Violeta , S̆egović Radmila
Published: July 2002

Amino acids in rat systemic and portal vein plasma and jejunal and ileal gut contents after 7 days of feeding normoprotein (NP) and protein‐free diet (PF) are investigated. Amino acid analyses revealed that ingestion of PF diet resulted in unusual amino acid patterns in both plasmas. Thus, while the levels and/or molar ratios of all indispensable amino acids were significantly decreased, those of several gluconeogenetic amino acids, especially of glycine and alanine, were increased in both plasmas, but particularly in portal. By contrast, the molar ratios of the majority of amino acids in jejunal and ileal contents were not changed by PF diet.
ADP‐ribosylation of specific membrane proteins in pheochromocytoma and primary‐cultured brain cells by botulinum neurotoxins type C and D
Authors: Matsuoka Ichiro , Syuto Bunei , Kurihara Kenzo , Kubo Shuichiro
Published: July 2002

Type C1 and D toxins produced by Clostridium botulinum caused ADP‐ribosylation of a protein of 24 kDa in membrane preparations of rat clonal pheochromocytoma cells (PC12) and of proteins of 25 and 26 kDa in neuron‐rich culture of fetal rat brain cells. The ADP‐ribosylation reaction was dependent on the presence of MgCl2, GTP and GTPγS. The results obtained suggested that the ADP‐ribosylation reaction is responsible for the development of the biological activity of the botulinum neurotoxins and that the target of this reaction may be novel GTP‐binding proteins localized on cell membranes.
UDP‐L‐arabinose‐hydroxyproline‐O‐glycosyltransferases in Volvox carteri
Authors: Günther Roland , Bause Ernst , Jaenicke Lothar
Published: July 2002

Hydroxyproline‐containing peptides of different length and amino acid sequence have been used to demonstrate UDP‐L‐arabinose‐hydroxyproline‐O‐glycosyltransferases in a crude microsomal fraction from the green alga Volvox carteri. The formation of O‐glycosidic linkages by transfer of UDP‐activated arabinose to the side chain of hydroxyproline was concluded from the resistance of the glycopeptides under the basic conditions of β‐elimination and their susceptibility to hydrolysis by trifluoroacetic acid. This treatment yielded arabinose as the only cleavage product. Arabinose transfer to the various peptide substrates was found to be stimulated by low concentrations of detergent, to require divalent cations and to proceed optimally at pH values around 7.0. The smallest arabinose acceptor peptide was the tripeptide Tyr‐Hyp‐Lys. The glycosyl acceptor electivity increased with increasing numbers of repeated hydroxyproline residues, suggesting that hydroxyproline clusters critically affect substrate recognition by the Volvox transferase(s).
The fluorescence intensity of the lipophilic probe N‐phenyl‐1‐naphthylamine responds to the oxidation‐reduction state of the respiratory chain in everted membrane vesicles of Escherichia coli
Authors: Sedgwick Edward G. , Bragg Philip D.
Published: July 2002

N‐Phenyl‐1‐naphthylamine (NPN), a reagent which has been used previously to probe the fluidity or microviscosity of the membrane lipids of intact cells of Escherichia coli, was found to respond to metabolic changes in everted inner membrane vesicles from this organism. NPN as bound to the vesicles to produce a steady‐state level of fluorescence intensity. Addition of substrate or ATP did not alter the fluorescence. However, following complete removal of oxygen from the medium by oxidation of substrate through the respiratory chain, there was an increase in the fluorescence of NPN. Reoxidation of the components of the respiratory chain by the addition of oxygen, ferricyanide, fumarate or nitrate decreased fluorescence to the steady‐state level until the oxidant had been completely reduced. The fluorescence changes were insensitive to the state of energization of the membrane. It is proposed that NPN responds to the state of reduction of components of the respiratory chain either directly by reacting with a component of the chain or indirectly through an effect transmitted to the membrane by a change in the conformation of respiratory chain components.
Somatostatin inhibits VIP‐ and isoproterenol‐stimulated cyclic AMP accumulation in rat prostatic epithelial cells
Authors: Carmena M.J. , Sancho J.I. , Fernández-Gonzalez M.A. , Escudero F. , Prieto J.C.
Published: July 2002

The dual regulation of cyclic AMP accumulation was studied in rat prostatic epithelial cells incubated with somatostatin, vasoactive intestinal peptide (VIP), and the β‐adrenergic agent isoproterenol. Somatostatin noncompetitively inhibited the stimulatory effect of VIP and isoproterenol, but it did not alter basal cyclic AMP levels. In addition to the multifactorial regulation of the cyclic AMP system in rat prostatic epithelium, these results suggest that somatostatin may play a physiological role at this level.
An enzyme catalysing the liberation of N‐acetylglucosamine from N‐acetylglucosaminyl pyrophosphorylpolyprenol in Bacillus polymyxa membranes
Authors: Murazumi Naoyuki , Yokoyama Kohei , Araki Yoshio , Ito Eiji
Published: July 2002

A novel enzyme which specifically hydrolyses N‐acetylglucosaminyl pyrophosphorylpolyprenol to liberate N‐acetylglucosamine was found in membranes of Bacillus polymyxa AHU 1385. The enzyme seems to be inactive toward α‐N‐acetylglucosaminyl phosphorylundecaprenol, β‐N‐acetylglucosaminyl phosphorylundecaprenol, N‐acetylglucosamine 1‐phosphate, N‐acetylglucosamine 1‐pyrophosphate, or UDP‐N‐acetyl‐glucosamine. Much lower activities of the same enzyme were also found in membranes of several other strains of Bacilli.
DNA‐dependent RNA polymerase from the chlorotetracycline producing strain of Streptomyces aureofaciens
Authors: Šimúth J. , Stembach H. , Zelinka J. , Chomutov R.M. , Nedospasov A.A.
Published: July 2002

RNA polymerase from Streptomyces aureofaciens has been purified by polyethyleneimine precipitation followed by chromatography first on DEAE‐cellulose, then heparin‐Sepharose and finally on an aminooxybutylcellulose matrix containing immobilised S. aureofaciens DNA. The enzyme is composed of three subunits of ∼ 145, 136 and 44 kDa that are in a ratio of approx. 1:1:2. In many isolations two additional subunits of ∼ 68 and 39 kDa and some minor protein bands of ∼ 110 85 and 61 kDa are also present. Thus, the structure of this enzyme is very similar to other bacterial RNA polymerases, exhibiting an α2ββ′ core and the additional proteins θ and σ.
Studies on rDNA from the extreme thermophilic eubacterium Thermus thermophilus HB8
Authors: Hartmann Roland K. , Vogel Detlef W. , Kröger Bernd , Ulbrich Norbert , Erdmann Volker A.
Published: July 2002

23 S ribosomal ribonucleic acid gene from the extreme thermophile eubacterium Thermus thermophilus HB8 has been cloned in pBR322, and the nucleotide sequence of region D has been determined, which encompasses 873 nucleotides at the 3′‐end of the RNA. We compare the primary and secondary structure of this region with the respective part of the 23 S rRNA from Escherichia coli and Bacillus stearothermophilus. A high level of structural conservation can be observed, throughout the RNA domain, albeit the usage of G/C basepairs is substantial even in comparison with another thermophilic eubacterium B. stearothermophilus. It is surprising that, in contrast to the usage of 3′U‐G5′, the occurrence of 3′G‐U5′ is comparable in E. coli as well as in B. stearothermophilus and T. thermophilus. Furthermore, it is most remarkable that the use of 3′A‐U5′ and 3′U‐A5′ is, compared to E. coli, only slightly reduced in B. stearothermophilus, but drastically decreased in T. thermophilus.
Energy transduction by the photosynthetic reaction center complex from Rhodopseudomonas viridis
Authors: Skulachev Vladimir P.
Published: July 2002

Comparison of data of (i) the high‐resolution X‐ray analysis of the Rhodopseudomonas viridis reaction center complex and (ii) direct (with a voltmeter) measurement of charge displacement in the same complex is presented. It is revealed that 95% of the energy stored in the form of the membrane electric potential is due to electron and 5% to proton transfers. Five stages of the overall process were found to contribute to the light‐induced membrane charging, namely electron transfers from excited bacteriochlorophyll dimer to bacteriopheophytin, from bacteriopheophytin to MQ, from heme c‐559 to bacteriochlorophyll dimer, from heme c‐556 to heme c‐559 and H+ transfer from cytoplasmic water to the bound CoQ. The contribution of each stage to the energy storage depends upon the dielectric constant value in the respective part of the complex rather than upon the distance covered by the electron or proton. It appears to be higher, the deeper the redox groups involved are immersed into the membrane.
Binding of heparin or dermatan sulfate to thrombin is essential for the sulfated polysaccharide‐accelerated inhibition of thrombin by heparin cofactor II
Authors: Yamagishi Ryoichi , Koide Takehiko , Sakuragawa Nobuo
Published: July 2002

Heparin cofactor II (HC II) and thrombin were chemically modified with pyridoxal 5′‐phosphate, and their effects on the inhibition of thrombin by HC II in the presence of heparin or dermatan sulfate were studied. The inhibition of thrombin by HC II was enhanced about 7000‐fold in the presence of heparin or dermatan sulfate. However, this enhancement by heparin dwindled to 110‐ and 9.6‐fold when the modified HC II and the modified thrombin, respectively, were substituted for native proteins. Essentially identical results were obtained from the experiments using dermatan sulfate. These results indicate that the binding of heparin or dermatan sulfate to both thrombin and HC II is required for the sulfated polysaccharide‐dependent acceleration of the thrombin inhibition by HC II, and the binding to thrombin is more essential for the reaction.