bluuebunny/crossref_metadata_2025_split huggingface dataset:
Evidence that 2‐allyl‐2‐isopropylacetamide, phenobarbital and 3,5‐diethoxycarbonyl‐1,4‐dihydrocollidine induce the same cytochrome P450 mRNA in chick embryo liver
Authors: BROOKER John D. , SRIVASTAVA Gopesh , BORTHWICK Iain A. , MAY Brian K. , ELLIOTT William H. 
Published: March 2005

The induction of cytochrome P450 in chick embryo liver has been studied using three different porphyrinogenic drugs, 2‐allyl‐2‐isopropylacetamide, 3,5‐diethoxycarbonyl‐1,4‐dihydrocollidine and phenobarbital. Pulse‐labelling studies have shown that for each drug the cytochrome P450 synthesised either in ovo or in a wheat germ translation system reacted immunologically with antibody raised against the purified 2‐allyl‐2‐isopropylacetamide‐induced enzyme (Mr= 50 000).To investigate whether this is due to the three drugs inducing the same protein or different proteins with common immunological determinants, nucleic acid hybridization studies have been carried out using a recently characterised 2‐allyl‐2‐isopropylacetamide‐induced cytochrome P450 cloned cDNA probe [Brooker, J. D. et al. (1982) Eur. J. Biochem. 129, 325–333]. It has been shown that the mRNA induced by each drug hybridizes with this probe and all are of similar size. The melting profile of the mRNA · cDNA hybrids indicates that the mRNAs induced by the three drugs have at least 98% homology with the cDNA probe.Restriction endonuclease digestions of total chick embryo genomal DNA and a chick cytochrome P450 genomal clone indicates that the cytochrome P450 gene homologous with the cDNA probe is represented in the genome only once.These results strongly suggest that the three drugs cause increased levels of the same cytochrome P450 mRNA, possibly due to enhanced expression of the same gene.Results are also presented which show that other cytochrome‐P450‐inducing drugs, 3‐methylcholanthrene, β‐naphthoflavone or pregnenolone‐16 α‐carbonitrile do not increase the level of the 2‐allyl‐2‐isopropylacetamide‐inducible mRNA but rather reduce it to a level which was lower than that of the untreated controls.
p‐HMW‐Collagen, a minor collagen obtained from chick embryo cartilage without proteolytic treatment of the tissue
Authors: BRUCKNER Peter , MAYNE Richard , TUDERMAN Leena 
Published: March 2005

The fragments of minor collagens of cartilages, called HMW and LMW, were isolated after pepsin treatment of sternal cartilages of young chickens and were shown to be entirely triple‐helical molecules as judged by their circular dichroic spectra. Studies on renaturation kinetics of HMW suggested that the interchain disulfide bonds in HMW reside at one of the ends of the so‐called long arm.Polyclonal antibodies against HMW were raised and affinity purified. These antibodies did not cross‐react with type II collagen nor with other minor collagens such as LMW and 1α, 2α, 3α collagen in native or denatured structure.The antibodies were used to identify HMW‐related molecules which were synthesized by embryonic chick cartilages in vitro. Some of these molecules were secreted into the organ culture medium and could be recovered from it by ammonium sulfate precipitation. Polyacrylamide gel electrophoresis of this precipitate gave one band of high molecular weight which could be reduced to two bands migrating slightly faster than the α1(II) chain when identified by immunoblotting. These bands could also be identified among about six radiolabelled polypeptides present in the ammonium sulfate precipitate of medium proteins when analysed by polyacrylamide gel electrophoresis followed by fluorography.The same polypeptides could be recovered from the medium by immunoprecipitation with anti‐HMW antibodies. Their presence in cartilage tissue was shown by immunoblotting of material extracted from cartilage tissue and separated on polyacrylamide gels. We suggest that the protein containing these polypeptide chains represents the parent molecule of the peptic fragment HMW as it is synthesized in vivo and have designated it p‐HMW‐collagen.
Carnitine palmitoyltransferase I
Authors: SIRE Oliver , MANGENEY Marise , MONTAGNE Jaqueline , NORDMANN Roger , NORDMANN Joseph 
Published: March 2005

Palmitate oxidation by liver mitochondria from rats treated with d‐galactosamine (GalN) was markedly inhibited, 3 h after administration.

The mitochondrial defect responsible for this inhibition was shown to be an inhibition of the activity of palmitoylcarnitine transferase I (EC 2.3.1.21).
Apparent Km of the enzyme remained unchanged whereas apparent V was reduced by 30%.
Addition of 10 mM GalN did not impair the activity of palmitoylcarnitine transferase I in mitochondria isolated from normal rats.
Inhibition of palmitoylcarnitine biosynthesis by GalN treatment was completely reversed by phospholipid supply.
At this stage of intoxication, mitochondrial phospholipid content was decreased whereas incorporation of [14C]palmitate into phospholipids in isolated hepatocytes was drastically inhibited; the phosphatidyl‐choline/phosphatidylethanolamine ratio was reduced by 33%.

The results obtained from these studies show that the depletion of the phospholipid membrane content could account for the altered functional activity of palmitoylcarnitine transferase I.
Cholinergic synaptic vesicles from Torpedo marmorata contain an atractyloside‐binding protein related to the mitochondrial ADP/ATP carrier
Authors: STADLER Herbert , FENWICK Elizabeth May 
Published: March 2005

Atractyloside is known to bind to the ADP/ATP translocase of the inner mitochondrial membrane, a complex formed by two basic protein subunits of relative molecular mass around 30000. We found that synaptic vesicles from the electric organ of Torpedo marmorata, which store acetylcholine and ATP, bind atractyloside as well. Similarly to mitochondria, a protein‐atractyloside complex could be solubilized from vesicle membranes with Triton X‐100. Characterization of the complex by gel filtration, isoelectric focusing and gel electrophoresis revealed that atractyloside was bound to protein V11, earlier described as a major vesicle membrane component with a relative molecular mass around 34000 and a basic isoelectric point.Since earlier experiments have already shown that uptake of ATP into isolated vesicles in vitro is inhibited by atractyloside, we can conclude now that V11 constitutes the nucleotide carrier of this secretory organelle. The structural and functional relationship of the mitochondrial and vesicular nucleotide translocases suggest a common evolutionary origin.
Subcellular fractionation of porcine neutrophils by nitrogen cavitation and sucrose‐density‐gradient centrifugation
Authors: CHIBBER Ramesh , CASTLE Alan G. 
Published: March 2005

Neutrophils, isolated in large quantities from porcine blood were disrupted by nitrogen cavitation and separated by differential centrifugation into a nuclear fraction and a post‐nuclear supernatant. The latter was subfractionated by sucrose density gradient centrifugation into cytosol, a fraction consisting of membrane vesicles and two granule‐rich fractions.
The membrane fraction accounted for 1.9% of the protein in the post‐nuclear supernatant, the light granule fraction for 2.2% and the dense granule fraction for 4.2%.
Catalase, lactate dehydrogenase and malate dehydrogenase were largely confined to the cytosol. The dense granule fraction contained the highest quantities of the hydrolytic enzymes, although the membrane fraction was also rich in alkaline and acid phosphatase and γ‐glutamyl transpeptidase activities.
Electron microscopy of the membrane fraction showed intact membrane vesicles, whereas the granular fractions consisted of electron‐dense, membrane‐bound granules. Two granular fractions were isolated which contained granules of differing size and density.
3H‐labeled wheat germ agglutinin bound to the surface of intact neutrophils and when these were disrupted and fractionated the membrane fraction showed a specific binding activity 16‐times greater than that of the cavitated sample.
The membrane fraction interacted with the detergent digitonin and as a result underwent density perturbation increasing from 1.13 g . cm−3 to 1.18 g . cm−3.
Dodecylsulphate‐polyacrylamide gel electrophoresis showed the membrane fraction to consist of at least 40 protein bands, with relative molecular masses ranging from 200000–16000. The granule fractions contained less protein bands, with a protein composition quite distinct from that of the membrane fraction.
Structure of the Mo‐Fe protein component of Azotobacter vinelandii nitrogenase
Authors: VOORDOUW Gerrit , HAAKER Huub , VAN BREEMEN Jan F. L. , VAN BRUGGEN Ernst F. J. , EADY Robert R. 
Published: March 2005

The Mo‐Fe protein of nitrogenase from both Azotobacter vinelandii and Klebsiella pneumoniae (Av1 and Kp1. respectively) consists of four subunits of similar, but not identical, relative molecular mass. The hydrodynamic properties of Av1 (sedimentation and diffusion coefficient) and its total relative molecular mass are very similar to those of Kp1 and catalase from bovine liver, a tetramer of four identical subunits.By electron microscopy the Av1, Kp1 and catalase tetramers are seen as protein particles of diameter 9.0–10.0 nm; no details of the subunit structure can be observed. Av1 (but not Kp1) forms regular polymers of variable length at low ionic strength in the presence of MgCl2. The structure of these polymers, of diameter 21.2 nm, is complex. Optical diffraction studies give a smallest repeating distance of 8.4 nm (corresponding to the diameter of the Av1 tetramer) and indicate a four‐start helix. The latter structure is incompatible with a flat, square subunit arrangement of the Av1 tetramer as proposed by Stasny et al. [(1974) J. Cell. Biol. 60, 311–316]. We propose, therefore, that the subunit arrangement of the Av1 tetramer is of the tetrahedral type. This has also been proposed for the catalase tetramer from optical diffraction studies of electron micrographs of catalase tubes indicating a 222 symmetry [Kiselev, D. A., De Rosier, N. J. and Klug, A. (1968) J. Mol. Biol. 35, 561–566]. Our proposal is in agreement with the recent finding that Av1 protein crystals belong to the P21 space group [Weiniger, M. S. and Mortenson, L. E. (1982) Proc. Natl Acad Sci. USA, 79, 378–380].
Properties of the chromatin repair activity against O6‐ethylguanine lesions in DNA
Authors: RENARD André , VERLY Walter G. , MEHTA Jitendra R. , LUDLUM David B. 
Published: March 2005

Chromatin proteins from rat liver contain a repair activity that removes O6‐ethylguanine from ethylnitrosoureatreated DNA. This activity does not depend on divalent cations and works in the presence of EDTA, but does depend on the presence of free thiol groups. Thus, it is destroyed by N‐ethylmaleimide and is protected by dithiothreitol. The repair activity on single‐stranded DNA is only 20% of what it is on double‐stranded DNA; its half‐life at 35°C is 55 min, but DNA, ethylated or not, affords some protection.The repair reaction is a transethylation from O6‐ethylguanine in DNA onto two different cysteine residues contained in acceptor proteins. The reaction can be followed by monitoring the appearance of ethylated proteins or by disappearance of O6‐ethylguanine from DNA.
Interaction between photoexcited rhodopsin and peripheral enzymes in frog retinal rods
Authors: PFISTER Claude , KÜHN Hermann , CHABRE Marc 
Published: March 2005

The major peripheral and soluble proteins in frog rod outer segment preparations, and their interactions with photoexcited rhodopsin, have been compared to those in cattle rod outer segments and found to be similar in both systems. In particular the GTP‐binding protein (G) has the same subunit composition, the same abundance relative to rhodopsin (1/10) and it undergoes the same light and nucleotide‐dependent interactions with rhodopsin in both preparations.Previous work on cattle rod outer segments has shown that photoexcited rhodopsin (R*), in a state identified with metarhodopsin II, associates with the G protein as a first step to the light‐activated GDP/GTP exchange on G. The complex R*‐G is stable in absence of GTP, but is rapidly dissociated by GTP owing to the GDP/GTP exchange reaction. Low bleaching extents (< 10% R*) in absence of GTP therefore create predominantly R*‐G complexes, whereas bleaching in presence of GTP creates free R*. We report here that, under conditions of complexed R*, two reactions of R* in frog rod outer segments are highly perturbed as compared to free R*: (a) the spectral decay of metarhodopsin II (MII) into later photoproducts, and (b) the phosphorylation of R* by an ATP‐dependent protein kinase.a) The spectral measurements have been performed using linear dichroism on oriented frog rod outer segments; this technique allows discrimination between MII and later photoproducts absorbing at the same wavelength. Association of R* with G leads to a strong reduction of the amount of MIII formed and to an acceleration of the decay of MIII. Furthermore, MII is significantly stabilized, in agreement with the hypothesis that MII is the intermediate which binds to G.b) The phosphorylation of R* is strongly inhibited under conditions of R*‐G complex formation as compared to free R*.Interferences between reactions at the three sites involved in R* are discussed: the retinal binding site in the hydrophobic core is sensitive to the presence of GTP‐binding protein at its binding site on the cytoplasmic surface of R*: the kinase and the GTP‐binding protein compete for access to their respective binding sites, both located on the surface of R*.We also observed a slow and nucleotide‐dependent light‐induced binding of a protein of molecular weight 50 000, which we consider as the equivalent of the 48 000 Mr light‐dependent protein previously identified in cattle rod outer segments.
Minactivin: A human monocyte product which specifically inactivates urokinase‐type plasminogen activators
Authors: GOLDER Jeffrey P. , STEPHENS Ross W. 
Published: March 2005

Culture supernatants from monolayers of human peripheral monocytes strongly inhibited colorimetric assays of urokinase in which plasmin was measured by esterolysis. This inhibitory activity of monocyte culture supernatant was enhanced after culture with muramyl dipeptide. Inhibition was specific for plasminogen activators of Mr 52 000 and 36 000, as shown by three methods: (1) inhibition of plasminogen‐dependent fibrinolysis; (2) inhibition at the level of plasminogen activation in a colorimetric assay; (3) the irreversible loss of plasminogen‐activating activity, as evidenced by electrophoresis, after preincubation with culture media. The factor responsible for this inactivation (which we propose to call minactivin) had an apparent Mr of 66 000 on Sephacryl S 300 gel chromatography and interacted with enzyme in a biphasic manner: a rapid partial inhibition (reversible by sodium dodecyl sulphate) was followed by slow inactivation (irreversible by sodium dodecyl sulphate).It is proposed that secretion of minactivin by monocytes may contribute to regulation of extracellular proteolysis at sites of tissue injury.
Ornithine‐containing lipid of Bordetella pertussis, a new type of hemagglutinin
Authors: KAWAI Yohko , YANO Ikuya 
Published: March 2005

The ornithine‐containing lipids of six strains (phases I–IV) of Bordetella pertussis were prepared from the total extractable cellular lipids by thin‐layer chromatography and treatment with phospholipase A. They were compared with those prepared from two strains each of Bordetella parapertussis and Bordetella bronchiseptica. The structures of the ornithine‐containing lipid of B. pertussis and the other two species were resolved by acid and alkaline hydrolysis, gas‐liquid chromatography, infrared absorption spectroscopy, amino acid analysis and combined gas‐liquid chromatography/mass spectrometry. The main structure of the aminolipid of the three species of Bordetella was 3‐hydroxyhexadecanoic acid, amide‐linked to ornithine and esterified to the second hexadecanoic acid. The aminolipid of B. pertussis Sakurayashiki (phase III) exhibited high hemagglutinating activity for human and rabbit erythrocytes, having a minimum hemagglutinating concentration of 1 γg/ml against 8–16 γg/ml for the other strains of Bordetella.All of these aminolipids showed some degree of microheterogeneity. Because the 3‐hydroxyhexadecanoic acid content was especially high in strain Sakurayashiki, it was presumed that the intensity of hemagglutinating activity of the aminolipid was affected by the chain length of the central 3‐hydroxy fatty acid, that is the aminolipid containing 3‐hydroxyhexadecanoic acid had high hemagglutinating activity. The hemagglutination was inhibited by phosphatidylcholine at concentrations of more than 20 γg/ml. Other inhibitory substances were cysteine, sphingomyelin, acidic amino acids, histidine, unsaturated fatty acid and basic amino acids. Furthermore, the divalent cations Ca2+ and Mg2+ inhibited this hemagglutination at a concentration of 1 mM. The O‐deacylated ornithine‐containing lipid that had lost hexadecanoic acid did not have any hemagglutinating activity but did have hemolytic activity. Observation by electron microscopy indicated that erythrocytes were combined by the liposomes of the ornithine‐containing lipids.On the basis of these results, the proposed mechanism of hemagglutination by the aminolipids is that the liposomes of the aminolipids combine erythrocytes by hydrophobic interaction between the fatty acid moieties of the aminolipid and the lipids of the surface of erythrocytes, and by ionic interaction between the ornithine of the aminolipid and the protein of the surface of the erythrocytes. In addition, the hemagglutinating activity of phosphatidylserine was found to be due to its similar structure to that of the ornithine‐containing lipid and the mechanism was also presumed to be similar. The mechanism of hemagglutination by these aminolipids was distinct from that of lectins.
Mutual conformational changes of tryptophanyl‐tRNA synthetase and tRNATrp in the course of their specific interaction
Authors: BERESTEN Sergey , SCHEINKER Vladimir , FAVOROVA Olga , KISSELEV Lev 
Published: March 2005

tRNATrp (beef, yeast) is capable of accelerating limited tryptic hydrolysis of the N‐terminal part in the polypeptide chains of dimeric beef pancreas tryptophanyl‐tRNA synthetase; it can also eliminate the protective effect of tryptophanyl adenylate on the enzyme proteolysis. The effect of tRNA on the proteolysis is manifested even when the 3′‐CCA terminus is removed. It has been concluded that the conformation of the synthetase changes when it forms a complex with tRNATrp. Yeast tRNATrp lacking the 3′‐half of the acceptor stem can still interact with the synthetase and, to certain extent, induces changes in the conformation of the latter.The susceptibility of single‐stranded and double‐stranded regions of tRNATrp to cleavage with endonucleases has been studied, and the results are indicative of the fact that, regardless of considerable differences in the nucleotide sequence of yeast and beef tRNATrp, their three‐dimensional structures are similar. This fact is consistent with the finding that parameters for the interaction of these tRNAsTrp with beef tryptophanyl‐tRNA synthetase are rather close.The three‐dimensional structure of tRNATrp is altered when the enzyme forms a complex with it, as seen from (a) a change in the circular dichroic spectrum and (b) an elevated susceptibility of the anticodon and, apparently, acceptor stems to cleavage with nuclease.The conversion of exposed cytidine residues in tRNATrp into uridine residues results in a loss of the acceptor activity; the capability to accelerate limited tryptic hydrolysis of tryptophanyl‐tRNA synthetase is also lost although the enzyme‐substrate complex, as seen from circular dichroic spectra, can still be formed. The conversion of cytosine in the anticodon stem into uracil modifies the conformation of the anticodon stem.The anticodon arm (including the anticodon) and the acceptor stem play an essential role in the interaction between tRNATrp and tryptophanyl‐tRNA synthetase.
Novel approach to the study of the regulation of hormone‐sensitive lipase in rat adipocytes
Authors: MOONEY Robert A. , EBERSOHL Richard D. , McDONALD Jay M. 
Published: March 2005

Use of a digitonin‐permeabilized rat adipocyte preparation overcomes inherent problems which occur when currently used broken cell systems are utilized for studying the regulation of hormone‐sensitive lipase. The effect of digitonin on plasma membrane permeability was concentration‐dependent being nearly maximum at 20 μg/ml as assessed by (a) leakage of 85% cellular lactate dehydrogenase after 30 min, (b) the efflux of 72% preloaded cellular (86Rb) rubidium within 10 min and (c) immediate inhibition of glucose oxidation.Hormone‐modulated rates of lipolysis were preserved in this preparation. Following maximal activation of lipolysis in adipocytes with catecholamines, the rate of lipolysis in intact cells and digitonin‐treated cells was elevated 26‐fold and 20‐fold respectively, while the rate in homogenates from these cells was elevated only 2.8‐fold. Insulin suppressed catecholamine‐dependent activation of lipolysis by at least 90% when subsequently measured in intact cells and digitonin‐treated cells. Insulin suppression was only 56% when measured in homogenates.The hormone‐sensitive lipase in permeabilized cells, as opposed to intact cells and homogenates, was activated by cyclic AMP to a degree that approached activation by catecholamines. In homogenates, cyclic AMP (1.0 mM) plus ATP (0.25 mM) activated the lipase only 36%, while neither alone had any effect. In digitonin‐permeabilized cells, however, exogenous cyclic AMP alone activated lipolysis in a concentration‐dependent manner with 1 μM, 30 μM and 1.0 mM cyclic AMP activating lipolysis by 41%, 250% and 1300% respectively. In contrast, lipolysis in intact cells was activated by 0%, 25% and 250% by 1 μM, 30 μM and 1.0 mM cyclic AMP. Also in digition‐treated preparations, ATP alone activated lipolysis 40%, but ATP plus cyclic AMP activated lipolysis to only 74% of the level due to cyclic AMP alone.These studies indicate that the permeabilized adipocyte preparation is an excellent system for investigating the mechanism of regulation of the hormone‐sensitive lipase by permitting manipulation of the intracellular environment while preserving the physiological response of the lipase.
NAD+ analogue binding to glyceraldehyde‐3‐phosphate dehydrogenase
Authors: WALLÉN Leif , BRANLANT Guy 
Published: March 2005

A series of NAD+ analogues, modified on the pyridinium ring, have been tested for their enzymic properties in reactions with d‐glyceraldehyde‐3‐phosphate dehydrogenase form sturgeon muscle, rabbit muscle and Bacillus stearothermophilus. The observed activity, inhibition and binding data are correlated to the structure of the enzyme and coenzyme analogue by model building on a Vector General interactive graphic display system using coordinates from the B. stearothermophilus holoenzyme structure.Most of the analogues with substituents in the pyridinium‐3 position could be bound to glyceraldehyde‐3‐phosphate dehydrogenase, either in manner similar to NAD+ or in a completely different way with the substituted pyridinium ring rotated 110° or more around the glycosidic bond. This indicates different possible modes of binding of NAD+ analogues within the pyridinium binding subsite.Analogues with substituents in the pyridinium‐4 position are shown to be weakly bound to glyceraldehyde‐3‐phosphate dehydrogenase. This is explained by a strong interaction of the substituent in the 4 position with the residues Asn‐313 and Cys‐149.