bluuebunny/crossref_metadata_2025_split huggingface dataset:
Studies on uroporphyrinogen decarboxylase of etiolated Euglena gracilis Z
Authors: JUKNAT Adela Ana , SEUBERT Andreas , SEUBERT Sigrid , IPPEN Hellmut 
Published: March 2005

A 423‐fold purified fraction of uroporphyrinogen decarboxylase (EC 4.1.1.37) showing a specific activity of 770 units/mg protein has been employed in order to study some properties in etiolated Euglena gracilis Z.
Uroporphyrinogen decarboxylase has a relative molecular mass of 54000, an optimum pH of 7.2 and exhibits Michaelis‐Menten kinetics, employing both uroporphyrinogen I and uroporphyrinogen III as substrates.
Anaerobic conditions seem not to be necessary for uroporphyrinogen decarboxylase activity. Neither EDTA nor cysteine affected enzyme activity, whereas dithiothreitol produced a remarkable activation of coproporphyrinogen formation.
Kinetic data employing both substrates showed an accumulation of porphyrinogen (i.e. hexa‐ and heptaporphyrin) containing six or seven COOH groups, depending on the uroporphyrinogen concentration used.
An unusual elution profile of the intermediates on Sephacryl S‐200 was found.
Enzyme Nomenclature. Recommendations 1984
Authors: WEBB Edwin C. 
Published: March 2005

This document contains 275 entries for new enzymes which have come to light since the finalization of the 1984 edition at the end of November, 1983, and the publication of Supplement 1 (Eur. J. Biochem. 157, 1–26 (1986). The 51 entries marked with an asterisk are revisions of the corresponding entries in Enzyme Nomenclature (1984), published by Academic Press, New York.References containing a letter (e.g. 123a) are listed at the end of this article; numbers without letters refer to the Reference List in Enzyme Nomenclature (1984).Comments and suggestions may be sent to any member of NC‐IUB or JCBN: J. F. G. Vliegenthart (Chairman), J. R. Bull, C. R. Cantor, M. A. Chester, C. Liébecq (as representative of the IUB Committee of Editors of Biochemical Journals), G. P. Moss, J. Reedijk, W. Saenger, N. Sharon, or P. Venetianer, to its Secretary, A. Cornish‐Bowden, CNRS‐CBM2, 31 Chemin Joseph‐Aiguier, B. P. 71, 13402 Marseille Cedex 9, France or to E. C. Webb, 1/221 Kings Road, Mundingburra, Townsville, Australia, 4812.
Defective processing and binding of low‐density lipoprotein receptors in fibroblasts from a familial hypercholesterolaemic subject
Authors: KNIGHT Brian L. , GAVIGAN Sean J. P. , SOUTAR Anne K. , PATEL Dilip D. 
Published: March 2005

The properties of the low‐density lipoprotein (LDL) receptor were studied in skin fibroblasts from a homozygous familial hypercholesterolaemic subject, MM (MM cells), who exhibits a defect in the processing of the precursor form of the receptor. Despite the prolonged half‐life of the precursor (3 h), essentially all was eventually processed to the mature form, which was degraded with a half‐life of approximately 6 h. The receptor content of the MM cells, determined by radioimmunoassay, was slightly lower than normal and by immunoblotting it was estimated that 55% was present as the mature form at equilibrium. The mature receptor reached the cell surface and was internalized and recycled apparently normally, with about 60% accessible to pronase at any time. Immunoblotting, immunoassay and surface labelling with 125I all indicated that the cells contained twice as much surface receptor protein as would be expected from the binding of LDL.Maximum heparin‐releasable LDL binding to MM cells was 20% of normal and the apparent affinity for LDL was reduced. The cells did not show the marked increase in affinity and reduction in binding exhibited by normal cells when cooled to 4°C. Also, neither the apparent affinity of the receptors nor the maximum binding at 37 C was greatly affected by an antibody that has been shown to reduce the affinity and halve the binding of normal cells. The results suggest that the mutation in the LDL receptor gene of these cells affected the ability of the receptors to bind LDL in a similar manner to the antibody, possibly by promoting aggregation of receptors on the cell surface, and halving the amount of LDL bound.
Specificity of antibodies to sea anemone toxin III and immunogenicity of the pharmacological site of anemone and scorpion toxins
Authors: BAHRAOUI El Mostafa , AYEB Mohamed EL , GRANIER Claude , BERESS Laslo , ROCHAT Hervé 
Published: March 2005

Toxin III (ATX III) of the sea anemone (Anemonia sulcata) is a polypeptide containing 27 amino acid residues.It has no sequence similarity with other toxins (ATX I and II) from the same species, or with scorpion toxins, although they apparently act in a similar manner by prolonging action potentials. The specificity of ATX III antibodies was characterized using ATX III, ATX I, native and chemically modified ATX II, and scorpion α‐toxins. The results obtained suggest that a regior of ATX III, partially or totally overlapping the pharmacological site shared with ATX I and ATX II, is immunogenic. It includes a guanidino and at least two carboxylate groups. The corresponding region is not immunogenic in ATX I and ATX II. Anti‐(ATX III) antibodies recognize the similar regions of ATX I and ATX II and apparently do not recognize scorpion toxins.
Correlation between enzyme activity and hinge‐bending domain displacement in 3‐phosphoglycerate kinase
Authors: SINEV Michael A. , RAZGULYAEV Oleg I. , VAS Mária , TIMCHENKO Alexander A. , PTITSYN Oleg B. 
Published: March 2005

Diffuse X‐ray‐scattering data give evidence for large‐scale structural change in pig muscle 3‐phosphoglycerate kinase upon substrate binding. Simultaneous binding of 3‐phosphoglycerate and MgATP either to the unmodified enzyme or to its active methylated derivative leads to about an 0.1‐nm decrease in radius of gyration. These data coincide well with the previous data for yeast 3‐phosphoglycerate kinase.When, instead of methylation, the two reactive thiol groups of pig muscle 3‐phosphoglycerate kinase are carboxamidomethylated, the enzyme becomes inactive and the radii of gyration of its ‘apo’ and ‘holo’ forms do not differ within limits of experimental error.Thus, a correlation exists between the activity of 3‐phosphoglycerate kinase and its substrate‐induced largescale conformational change. This correlation is a strong argument in favor of the functional importance of domain locking in the reaction catalyzed by 3‐phosphoglycerate kinase.
The kinetics of the thermal deactivation of transglutaminase from guinea‐pig liver
Authors: NURY Sylvie , MEUNIER Jean‐Claude , MOURANCHE Annette 
Published: March 2005

By incubating native (N) transglutaminase from guinea‐pig liver at various temperatures and assaying it at 25°C, two steps in the irreversible deactivation process to the denatured form (D) have been found. The fitting of the data to the equations of two possible models (the two‐steps model and the two‐isoenzymes model) is only compatible with the first one (N→X→D). It is shown that the structure of the active intermediate, X, depends on the deactivation temperature and on the thermal history of the enzyme. This may mean that transglutaminase exists in a large number of microstates. Surprisingly, the activation energy of deactivation is lower than that of activity (36.6 ± 3.4 against 47.2 ± 2.2 kJ · mol−1). By deactivating transglutaminase at a constant temperature (55°C) and assaying it at variable temperatures, the activation energy of the intermediate, (X55), has been determined to be 40.2 ± 5 kJ · mol−1, of the same order of magnitude as the native form.Among several agents assayed, only Ca2+ had a positive effect on the thermal stability of this enzyme. At 40°C, transglutaminase was quite stable in the presence of Ca2+ (in its absence, the half‐life was 65 min) and at 45°C, its thermostability had been considerably increased, the half‐life being raised from 47 min to 275 min.
Mechanism of action of cysteinyl leukotrienes on glucose and lactate balance and on flow in perfused rat liver
Authors: IWAI Masaru , JUNGERMANN Kurt 
Published: March 2005

Rat livers were perfused at constant pressure via the portal vein with media containing 5 mM glucose, 2mM lactate and 0.2 mM pyruvate.

Leukotrienes C4 and D4 enhanced glucose and lactate output and reduced perfusion flow to the same extent and with essentially identical kinetics. They both caused half‐maximal alterations (area under the curve) of carbohydrate metabolism at a concentration of about 1 nM and of flow at about 5 nM. The leukotrine‐C4/D4 antagonist CGP 35949 B inhibited the metabolic and hemodynamic effects of 5 nM leukotrienes C4 and D4 with the same efficiency, causing 50% inhibition at about 0.1 μM.
Leukotriene C4 elicited the same metabolic and hemodynamic alterations with the same kinetics as leukotriene D4 in livers from rats pretreated with the γ‐glutamyltransferase inhibitor, acivicin.
The calcium antagonist, nifedipine, at a concentration of 50 μM did not affect the metabolic and hemodynamic changes caused by 5 nM leukotriene D4. The smooth‐muscle relaxant, nitroprussiate, at a concentration of 10 μM reduced flow changes, without significantly affecting the metabolic alterations.
Leukotriene D4 not only reduced flow; it also caused an intrahepatic redistribution of flow, restricting some areas from perfusion. Thus leukotrienes increased glucose and lactate output directly in the accessible parenchyma and, in addition, indirectly by washout from restricted areas during their reopening upon termination of application.
The phospholipase A2 inhibitor, bromophenacyl bromide, but not the cyclooxygenase inhibitor, indomethacin, at a concentration of 20 μM reduced the metabolic and hemodynamic effects of 5 mM leukotriene D4.
Stimulation of the sympathetic hepatic nerves with 2‐ms rectangular pulses at 20 Hz and infusion of 1 μM noradrenaline increased glucose and lactate output and decreased flow, similar to 10 nM leukotrienes C4 and D4. The kinetics of the metabolic and hemodynamic changes caused by the leukotrienes differed, however, from those due to nerve stimulation and noradrenaline.
The leukotriene‐C4/D4 antagonist, CGP 35949 B, even at very high concentrations (20 μM) inhibited the metabolic and hemodynamic alterations caused by nerve stimulation or noradrenaline infusion only slightly and unspecifically.

The present results suggest that (a) leukotriene C4 and leukotriene D4 modulate hepatic metabolism and hemodynamics with equal kinetics and efficiencies specifically, via leukotriene C4/D4 receptors; (b) they act without being dependent upon calcium uptake via slow calcium channels, in part indirectly, by stimulating the formation of other phospholipase A2 produts and (c) they do not have a major role in the actions of sympathetic hepatic nerves and circulating noradrenaline. Metabolic regulation by leukotrienes, which are formed only in non‐parenchymal cells, constitutes another example of a complex intra‐organ cell—cell communication between non‐parenchymal and parenchymal cells.
Nuclear basic protein transition during sperm differentiation
Authors: CHAUVIÈRE Muriel , MARTINAGE Arlette , BRIAND Gilbert , SAUTIÈRE Pierre , CHEVAILLIER Philippe 
Published: March 2005

The remodeling of nucleoproteins during dog‐fish spermiogenesis involves two successive nuclear protein transitions: the first from somatic‐type histones to transition proteins during the nuclear elongation of spermatids and the second leading to protamine‐DNA association in mature spermatozoa. The chromatin of elongating spermatids contains two transition proteins called S1 and S2. The amino acid sequence of protein S1, a polypeptide of 87 residues was determined previously [Chauvière, M., Martinage, A., Briand, G., Sautière, P. & Chevaillier, Ph. (1987) Eur. J. Biochem. 169, 105–111]. In the present paper, we report the elucidation of the primary structure of the minor transition protein S2 established by automated Edman degradation of the protein and of its fragments generated by cleavage at methionine and aspartate residues. S2 contains 80 residues and has a molecular mass of 9726 Da. S2 is mainly characterized by a high content of basic amino acids mostly represented by lysine, a relatively high level of hydrophobic residue, the presence of six phosphorylatable residues and the lack of cysteine. Its amino acid sequence shows that the N‐terminal half is highly basic, while the acidic residues are located in the C‐terminal part of the protein where more diversity in amino acids is noticed.The two transition proteins S1 and S2 share striking structural similarities. Few but significative similarities have been detected with the mammalian transition protein TP1 [Kistler, W. S., Noyes, C., Hsu, R. & Heinrikson, R. L. (1975) J. Biol. Chem. 250, 1847–1853], suggesting similar functions for all these proteins in chromatin remodeling during sperm differentiation. By contrast, the two dog‐fish spermatid‐specific proteins are structurally unrelated to sperm protamines and cannot be considered as their precursors.
On the role of the termination factor RF‐2 and the 16S RNA in protein synthesis
Authors: LANG Anne , FRIEMERT Constanze , GASSEN Hans Günter 
Published: March 2005

In the translational termination step of protein synthesis the three termination codons UAA, UAG or UGA are recognized by so‐called release or termination factors. The release factor RF‐1 interacts with UAG and UAA whereas RF‐2 is specific for UGA and UAA. Two mechanisms concerning the termination event have been discussed so far: recognition of the termination codon by the protein in a tRNA‐like manner or double‐strand formation between the codon and the 3′ end of the 16S rRNA which is stabilized by the termination factor. Using equilibrium dialysis we show that 40% of the ribosomes can bind UGAA in an RF‐2‐dependent manner.The stability with the correct combination RF‐2—UGA is tenfold higher as compared to the wrong termination codon UAG. We confirm prior findings that the termination factor RF‐2 is bound to the A‐site of the ribosome. In addition to the ribosomal proteins L2, L10, L7/L12 and L20 of the large subunit and S6 and S18 of the small subunit, the 16S rRNA became labelled when radioactive UGA was crosslinked to the ribosome in the presence of RF‐2. Our data support a mechanism of termination in which a double strand between the termination codon and the 3′ end of the 16S rRNA is formed as the starting event. The resulting RNA‐RNA double strand in turn may be recognized and stabilized by the termination factor.
The gene encoding dihydrolipoyl transacetylase from Azotobacter vinelandii
Authors: HANEMAAIJER Roeland , WESTPHAL Adrie H. , BERG Axel , DONGEN Walter VAN , de KOK Arie , VEEGER Cees 
Published: March 2005

The gene encoding the dihydrolipoyl transacetylase (E2) component from Azotobacter vinelandii has been cloned in Escherichia coli. High expression of the gene was found when the cells were grown for more than 14 h. The E2 produced was partially active, varying 10 and 90% in different experiments. By limited proteolysis of the protein it was shown that the catalytic domain was incorrectly folded, caused by formation of intermolecular or intramolecular S‐S bridges. The enzyme was fully activated after unfolding in 2.5 M guanidine hydrochloride containing 2 mM dithiothreitol, followed by refolding by dialysis. Active E2 was isolated in a simple three‐step procedure. It possessed a specific activity in the same order as that found after isolation of E2 from purified pyruvate dehydrogenase complex from A. vinelandii. Active E2 comprises about 7% of the total soluble cellular protein in the E. coli clone.By genetic manipulation, deletion mutants of E2 were created, one encoding the lipoyl domain and the N‐terminal half of the pyruvate‐dehydrogenase (E1)‐ and lipoamide‐dehydrogenase (E3)‐binding domain, the other encoding the catalytic domain and the C‐terminal half of the E1‐ and E3‐binding domain. In E. coli expression of both mutants was observed.
Isolation of heparin‐binding growth factors from bovine, porcine and canine hearts
Authors: QUINKLER Wolfgang , MAASBERG Michael , BERNOTAT‐DANIELOWSKI Sabine , LÜTHE Norbert , SHARMA Hari S. , SCHAPER Wolfgang 
Published: March 2005

Fresh bovine, porcine and canine hearts were homogenized and mitogens for mesoderm‐derived cells were purified in three different steps. Extraction by two different ammonium sulfate precipitations was followed by cation‐exchange chromatography and by heparin‐Sepharose affinity chromatography. A heparin‐Sepharose fraction from heart (eluted at 1.1 M NaCl) increased mitotic activity in serum‐deprived cultures of porcine aortic endothelial and smooth muscle cells, and in human fibroblasts. This mitogenic activity is potentiated by heparin and inhibited by γ‐interferon. The heart mitogenic fraction showed one double peak on HPLC at A215 and one polypeptide‐band on SDS/PAGE. These peaks and bands were identical to those obtained from bovine brain. The heart acidic fibroblast growth factor (aFGF) showed a positive signal in Western blots using antibodies raised against brain aFGF. Gas‐phase amino acid sequencing established that the mitogens were identical to aFGF and the N‐terminally truncated aFGF. Extraction in the presence of a protease inhibitor (pepstatin A) produced a higher‐molecular mass form of aFGF with a blocked amino terminus.Another mitogen, eluted at 1.6 M NaCl from heparin‐Sepharose, reacted with polyclonal antiserum against human recombinant basic fibroblast growth factor (bFGF) and showed a 66% (12 from 18 amino acids determined by gas‐phase sequencing) similarity with bFGF. This polypeptide increased the mitotic activity of the same cell lines but was more potent than aFGF.
Metabolism of cysteinyl leukotrienes in non‐recirculating rat liver perfusion
Authors: WETTSTEIN Matthias , GEROK Wolfgang , HÄUSSINGER Dieter 
Published: March 2005

The uptake, metabolism and biliary excretion of the cysteinyl leukotrienes LTC4, LTD4 and LTE4, were studied in a non‐recirculating rat liver perfusion system at constant flow in both antegrade (from the portal to the caval vein) and retrograde (from the caval to the portal vein) perfusion directions. During a 5‐min infusion of [3H]LTC4,[3H]LTD4 and [3H]LTE4 (10 nmol/l each) in antegrade perfusions single‐pass extractions of radioactivity from the perfusate were 66%, 81% and 83%, respectively. Corresponding values for LTC4 and LTD4 in retrograde perfusions were 83% and 93%, respectively, indicating a more efficient uptake of cysteinyl leukotrienes in retrograde than in antegrade perfusions. The concentrations of unmetabolized leukotrienes in the effluent perfusate were 8–12% in antegrade and 2–4% in retrograde perfusions. [14C]Taurocholate extraction from the perfusate was inhibited by LTC4 by only 3%, suggesting that an opening of portal‐venous/hepatic‐venous shunts does not explain the effects of perfusion direction on hepatic LTC4 uptake.
Following infusion of [3H]LTC4 and [3H]LTD4, in the antegrade perfusion direction, about 80% and 87%, respectively, of the radiolabel taken up by the liver was excreted into bile. In retrograde perfusions, however, only 40% and 57%, respectively, was excreted into bile and the remainder was slowly redistributed into the perfusate, indicating that leukotrienes were taken up into a hepatic compartment with less effective biliary elimination or converted to metabolites escaping biliary excretion. The metabolite pattern found in bile was not affected by the direction of perfusion. Biliary products of LTC4 were polar metabolites (31–38%), LTD4 (27–30%), LTE4 (about 1%) and N‐acetyl‐LTE4 (3–4%) in addition to unmodified LTC4 (17–18%).
LTC4 was identified as a major metabolite of [3H]LTD4 in bile, amounting to about 20% of the total radioactivity excreted into bile. This is probably due to a γ‐glutamyltransferase‐catalyzed glutamyl transfer from glutathione in the biliary compartment, as demonstrated in in vitro experiments. The presence of sinusoidal γ‐glutamyltransferase activity in perfused rat liver was shown in experiments on the hydrolysis of infused γ‐glutamyl‐p‐nitroanilide. 90% inhibition of this enzyme activity by AT‐125 did not affect the metabolism of LTC4.
When [3H]LTE4 was infused in the antegrade perfusion direction, biliary metabolites comprised N‐acetyl‐LTE4 (24%) and polar components (60%).
The data show (a) that cysteinyl leukotrienes are effectively taken up and metabolized during a single liver passage, the metabolites being excreted into bile and released into perfusate; (b) zonal hepatocyte heterogeneities regarding leukotriene effector, uptake and biliary excretion sites are likely to exist; and (c) LTD4 in bile is converted to LTC4, suggesting also a biosynthetic function of canalicular or biliary γ‐glutamyltransferase, at least in the experimental system of perfused rat liver.
Hormone‐induced expression of the CHSl gene from Saccharomyces cerevisiae
Authors: APPELTAUER Ute , ACHSTETTER Tilman 
Published: March 2005

When MATa cells of Saccharomyces cerevisiae have been treated with the mating hormone α‐factor an increase in chitin synthase zymogen, as well as chitin content in the cell‐wall fraction, have been reported. With a DNA probe derived from the cloned CHSl gene that codes for chitin synthase I [Bulawa, C. E., Slater, M., Cabib, E., Au‐Young, J., Sburlati, A., Adair, W. L. and Robbins, P. (1986) Cell 46, 213–225] a Northern analysis was conducted of CHSl‐specific transcripts. α‐Factor‐treated MATa cells revealed more than sixfold elevated steady‐state levels of CHSl mRNA as compared to control cells. MATα cells responded the same way when treated with a‐factor although induction rate was somewhat smaller. After hormone application a rapid increase in CHSl mRNA levels could be observed that occurred also in the absence of ongoing protein synthesis.In order to minimize possible side effects of CHSl‐coding sequences on expression and mRNA stability a CHSl::SUC2 chimaeric gene was constructed where 730 bp of the CHSl promoter region (+ 20 bp of the coding region) were fused in frame to a fragment of the SUC2 coding region. The fusion protein exhibits invertase activity that has been used to monitor CHSl promoter activity. By analysis of shortened versions of the CHSl promoter a 94‐bp DNA fragment has been identified that confers hormone inducibility to the CHSl promoter. According to the published sequence of the CHSl gene, this fragment contains four repeats of a TGAAACA consensus sequence previously identified in the α‐factor‐inducible BARl promoter [Kronstad, J. W., Holly, J. A. and MacKay, V. L. (1987) Cell 50, 369–377]. This heptamer may represent the cis‐acting element involved in mating‐hormone‐mediated gene expression in yeast.
Purification and characterization of an oxygen‐stable carbon monoxide dehydrogenase of Methanothrix soehngenii
Authors: JETTEN Mike S. M. , STAMS Alfons J. M. , ZEHNDER Alexander J. B. 
Published: March 2005

Carbon monoxide dehydrogenase was purified to apparent homogeneity from Methanothrix soehngenii. In contrast with the carbon monoxide dehydrogenases from most other anaerobic bacteria, the purified enzyme of Methanothrix soehngenii was remarkably stable towards oxygen and it was only slightly inhibited by cyanide.The native molecular mass of the carbon monoxide dehydrogenase of Methanothrix soehngenii determined by gel filtration was 190 kDa. The enzyme is composed of subunits with molecular mass of 79.4 kDa and 19.4 kDa in an α2β2 oligomeric structure. The enzyme contains 1.9 ± 0.2 (n= 3) mol Ni/mol and 19 ± 3 (n= 3) mol Fe/mol and it constitutes 4% of the soluble cell protein. Analysis of enzyme kinetic properties revealed a Km of 0.7 mM for CO and of 65 μM for methyl viologen. At the optimum pH of 9.0 the Vmax was 140 μmol of CO oxidized min−1 mg protein−1. The enzyme showed a high degree of thermostability.