bluuebunny/crossref_metadata_2025 huggingface dataset:
o‐Quinone/quinone methide isomerase: A novel enzyme preventing the destruction of self‐matter by phenoloxidase‐generated quinones during immune response in insects
Authors: Saul Steven J. , Sugumaran Manickam
Published: July 2002

Melanization and encapsulation of invading foreign organisms observed during the immune response in insects is known to be due to the action of activated phenoloxidase. Phenoloxidase‐generated quinones are deposited either directly or after self‐polymerization on foreign objects accounting for the observed reactions. Since the reactions of quinones are nonenzymatic, they do not discriminate self from nonself and hence will also destroy self‐matter. In this report we present evidence for the presence of a novel quinone/quinone methide isomerase in the hemolymph of Sarcophaga bullata which destroys long‐lived quinones and hence acts to protect the self‐matter. Quinone methides, formed by the action of this enzyme on physiologically important quinones, being unstable undergo rapid hydration to form nontoxic metabolites.
Receptor and effector interactions of Gs Functional studies with antibodies to the αs carboxyl‐terminal decapeptide
Authors: Simonds William F. , Goldsmith Paul K. , Woodard Charles J. , Unson Cecilia G. , Spiegel Allen M.
Published: July 2002

Antibodies generated to a synthetic decapeptide, RMHLRQYELL, representing the carboxyl‐terminus of Gs‐α have been characterized in immunoblots and functional studies. This antibody, designated RM, reacts exclusively with a doublet of proteins of 52 and 45 kDa in immunoblots of bovine brain and wild‐type S49 murine lymphoma cell membranes. No such reactivity is seen in membranes from cyc
− S49 cells, which lack Gs. RM blocks receptor‐mediated activation of Gs and adenylyl cyclase in membranes from wild‐type S49 cells. RM could also immunoprecipitate adenylyl cyclase activity in detergent extracts from GTP[γ]S‐ or fluoride‐preactivated bovine brain membranes; thus binding of αs to effector and carboxyl‐terminal antibody was mutually compatible. Such experiments provide an approach for the elucidation of functionally relevant interactions of G‐proteins with receptors and electors in the membrane.
Primary structure of a new cysteine proteinase inhibitor from pig leucocytes
Authors: Ritonja Anka , Kopitar Majda , Jerala Roman , Turk Vito
Published: July 2002

The primary structure of a pig leucocyte cysteine proteinase inhibitor, also called cathelin, was determined. The sequence was obtained from analyses of peptides isolated from the chymotryptic, endoproteinase Lys‐C and protease V8 digests, and by analysis of the peptides derived from the hydrolysis of the aspartyl‐prolyl bond of the carboxymethylated inhibitor. The inhibitor consists of 96 residues. The N‐terminal residue of the inhibitor is pyrrolidonecarboxylic acid. The amino acid sequence of cathelin suggests the appearance of a new family of cysteine proteinase inhibitors.
The primary structure of rat ribosomal protein L18a
Authors: Aoyama Yuji , Chan Yuen-Ling , Meyuhas Oded , Wool Ira G.
Published: July 2002

The amino acid sequence of rat ribosomal protein L18a was deduced from the sequence of nucleotides in a recombinant cDNA. Ribosomal protein L18a contains 175 amino acids and has a molecular mass of 20 047 Da. Hybridization of the cDNA to digests of rat nuclear DNA and to a preparation of poly (A)+ mRNA suggests that there are 8–11 copies of the L18a gene and that the mRNA for the protein is about 700 nucleotides in length. Rat L18a is related to Schizosaccharomyces pombe L17 and perhaps to Halobacterium marismortui L19.
Expression of retinoic acid receptor genes in keratinizing front of skin
Authors: Noji Sumihare , Yamaai Tomoichiro , Koyama Eiki , Nohno Tsutomu , Fujimoto Wataru , Arata Jiro , Taniguchi Shigehiko
Published: July 2002

We found, by an in situ hybridization method with riboprobes synthesized from human cDNA of the retinoic acid receptor (RAR), that the RAR genes (predominantly γ‐subtype) are intensively expressed in the epidermis of normal and psoriasic human skins, and also in keratinizing fronts of 4‐day‐old mouse skins, nail matrices and hair follicles. Thus, target cells of retinoic acid in the skins are concluded to be keratinocytes, which is quite consistent with the fact that retinoic acid regulates keratinization of epidermis in vivo and also modulates expression of the keratin gene in vitro.
Translational active mRNPs from rabbit reticulocytes are qualitatively different from free mRNA in their translatability in cell‐free system
Authors: Minich Waldemar B. , Korneyeva Nadezhda L. , Ovchinnikov Lev P.
Published: July 2002

The translatability of polyribosomal and free mRNPs from rabbit reticulocytes and their mRNA was compared. Both classes of mRNPs turned out to be active in rabbit reticulocyte lysates. Considerable differences between mRNPs and mRNA have been revealed. The most striking feature of mRNPs was that high concentrations of mRNPs do not inhibit protein biosynthesis, whereas high concentrations of mRNA strongly inhibit this process. This inhibition is specific for mRNA and does not occur at the addition of the same amount of rRNA from E. coli. The features of mRNP translation are not the result of addition of the supplementary translation factors within particles. The specific function of mRNP proteins in the process of translation is under discussion.
A special repressor/activator system controls distribution of mRNA between translationally active and inactive mRNPs in rabbit reticulocytes
Authors: Minich Waldemar B. , Korneyeva Nadezhda L. , Berezin Yuri V. , Ovchinnikov Lev P.
Published: July 2002

Translation of free mRNPs and polyribosomal mRNPs from rabbit reticulocytes was studied in a rabbit reticulocyte and wheat germ cell‐free systems. It has been shown that the translation efficiency of polyribosomal mRNPs and the mRNA isolated from the particles is nearly the same in both systems. At the same time, free mRNPs' translatability, which is high in the homologous cell‐free system, is very low in the system from wheat germs. Translation efficiency of free mRNPs in the wheat germ system can be restored by addition of 0.5 M KCl‐wash of rabbit reticulocyte ribosomes. These results testify to the existence of some special repressor/activator system which controls the distribution of mRNA between free mRNPs and polyribosomes in rabbit reticulocytes.
Human embryonic myosin heavy chain cDNA
Authors: Eller Mark , Stedman Hansell H. , Sylvester James E. , Fertels Scott H. , Wu Qi-long , Raychowdhury Malay K. , Rubinstein Neal A. , Kelly Alan M. , Sarkar Satyapriya
Published: July 2002

Myosin heavy chain cDNA; Nucleotide sequence; Amino acid sequence; Myosin rod; Chromosomal mapping; Gene transcription; (Human embryo)
Lipid lateral diffusion and membrane organization
Authors: Tocanne Jean-François , Dupou-Cézanne Laurence , Lopez André , Tournier Jean-François
Published: July 2002

Phospholipid; Photobleaching; Lateral diffusion; Membrane organization; Macrodomain; Microdomain
Regional expression of a MCD‐peptide and dendrotoxin I‐sensitive voltage‐dependent potassium channel in rat brain
Authors: Séquier Jean-Marc , Brennand John , Barhanin Jacques , Lazdunski Michel
Published: July 2002

In situ hybridization histochemistry has been used to analyze the regional expression of a class of voltage‐dependent K+ channel that is sensitive to two polypeptide toxins (MCD peptide and dendrotoxin I) that produce spectacular effects on brain function. A heterogeneous expression of this K+ channel was observed throughout the brain. High naRNA contents were observed in the granule cells of the gyrus dentatus as well as in pyramidal cells of the Ammon hom (CA3 > CA1) and in the cerebellum. Conversely, low levels of expression were found in basal ganglia (caudate putamen, globus pallidus, and ventral pallidum).
Disulfide pairings in geographutoxin I, a peptide neurotoxin from Conus geographus
Authors: Hidaka Yuji , Sato Kazuki , Nakamura Hideshi , Kobayashi Jun'ichi , Ohizumi Yasushi , Shimonishi Yasutsugu
Published: July 2002

The three intramolecular disulfide linkages of geographutoxin I, a peptidic neurotoxin isolated from the venom of the marine snail Conus geographus, were examined by a novel method for determination of the positions of disulfide linkages in peptides [(1989) Bull. Chem. Soc. Jp. 62, 1986–1994]. The disulfide bridges were found to be between Cys3 and Cys15, Cys4 and Cys20, and Cys10 and Cys21, indicating that geographutoxin I has a rigid conformation consisting of three loops stabilized by these three disulfide linkages.
Conservation of engrailed‐like homeobox sequences during vertebrate evolution
Authors: Holland Peter W.H. , Williams Nicola A.
Published: July 2002

The Drosophila melanogaster developmental gene engrailed (en) is a member of a distinct subfamily of homeobox genes with a wide phylogenetic distribution. Here we report the use of reduced stringency polymerase chain reaction (PCR) to amplify and clone 8 genes related to en from 5 vertebrate species, including representatives of the most ancient vertebrate lineages. Nucleotide and deduced amino acid sequence comparisons between mouse, toad, zebrafish, lamprey and hagfish genes reveal extensive evolutionary conservation, and suggests that 2 en‐like genes have been retained in most vertebrate lineages.
Chromatin structure of Drosophila melanogaster ribosomal genes
Authors: Belikov S.V. , Dzherbashyajan A.R. , Preobrazhenskaya O.V. , Karpov V.L. , Mirzabekov A.D.
Published: July 2002

The chromatin structure of ribosomal genes of D. melanogaster has been studied by crosslinking proteins to DNA. We found that a number of histone contacts with DNA through histidine in the approximately 1 kb‐long region surrounding the transcription initiation site, coding regions and the region of 240 bp‐long repeats from the intergenic spacers (Alu‐repeats) were weakened as compared to the inactive chromatin of the type II insertion. A protein with the molecular mass of 50 kDa (p50), associated with all DNA sequences analysed, has been discovered. Another protein with molecular mass of about 70 kDa (p70) has been found to be specific only for the Alu‐repeats.
Functional involvement of calcium in the homologous up‐regulation of the 1,25‐dihydroxyvitamin D3 receptor in osteoblast‐like cells
Authors: van Leeuwen J.P.T.M. , Birkenhäger J.C. , Buurman C.J. , Schilte J.P. , Pols H.A.P.
Published: July 2002

In several cell types 1,25‐dihydroxyvitamin D3 (1,25(OH)2D3) causes up‐regulation of its receptor. The present study demonstrates that in the osteoblast‐like cell line UMR 106 this up‐regulation is inhibited by two different calcium channel blockers (nitrendipine, verapamil). Also with chelating extracellular calcium (EGTA) and by inhibition of calcium release from intracellular stores (TMB‐8) comparable results were obtained. These findings indicate that calcium is functionally involved in this cellular response to the steroid hormone 1,25(OH)2D3. Moreover, data obtained with EGTA show that the 1,25(OH)2D3 receptor level is closely regulated by the extracellular calcium concentration.
A protein conformational change associated with the photoreduction of the primary and secondary quinones in the bacterial reaction center
Authors: Nabedryk E. , Bagley K.A. , Thibodeau D.L. , Bauscher M. , Mäntele W. , Breton J.
Published: July 2002

A comparison is made between the PQA → P+Q−
A and PQAQB → P+QAQ−
B transitions in Rps. viridis and Rb. sphaeroides reaction centers (RCs) by the use of light‐induced Fourier transform infrared (FTIR) difference spectroscopy. In Rb. sphaeroides RCs, we identify a signal at 1650 cm−1 which is present in the P+QA‐minus‐PQA spectrum and not in the P+QAQ−
B‐minus‐PQAQB spectrum. In contrast, this signal is present in both P+Q−
A‐minus‐PQ−
A and P+QAQ−
B‐minus‐PQAQB spectra of Rps. viridis RCs. These data are interpreted in terms of a conformational change of the protein backbone near QA (possible at the peptide C≡O of a conserved alanine residue in the QA pocket) and of the different bonding interactions of QB with the protein in the RC of the two species.
Prediction of domain organisation and secondary structure of thyroid peroxidase, a human autoantigen involved in destructive thyroiditis
Authors: Banga J.Paul , Mahadevan Daruka , Barton Geoffrey J. , Sutton Brian J. , Saldanha Jose W. , Odell Eddy , McGregor Alan M.
Published: July 2002

Organ specific autoimmune diseases are relatively common immunological disorders in man which include thyroid autoimmune disease, insulin‐dependent diabetes mellitus and myasthenia gravis. The target autoantigens in some of these diseases have recently been characterised. In thyroid autoimmune disease this includes the key enzyme, thyroid peroxidase (TPO), which is involved in the generation of thyroid hormone. Structural knowledge about autoantigens such as thyroid peroxidase will allow a greater understanding of the interaction between autoantigens and the aberrant immune response, and facilitate the development of strategies for antigen‐specific therapeutic manipulation. We report here a prediction of the secondary structure of thyroid peroxidase, together with the results of circular dichroic spectroscopy of a homologous purified enzyme. A combination of 3 secondary structure prediction programs has been used, following multiple sequence alignment, and TPO has been found to consist mainly of α‐helical conformation, with little β‐sheet present. This structure prediction, together with knowledge of the exon‐intron boundaries allows a model for the domain organisation of the TPO molecule to be proposed.
Antibodies against the cytoplasmic tail can differentiate between the quaternary forms of the Mr 46000 mannose 6‐phosphate receptor
Authors: Nadimpalli Siva Kumar , Schmidt Bernhard , von Figura Kurt , Hille Annette
Published: July 2002

An antiserum against a peptide of the cytoplasmic tail of the M
r 46000 mannose 6‐phosphate receptor is described which recognizes preferentially the tetrameric versus the dimeric form of this receptor. This indicates that the conformation of the cytoplasmic tail, which harbours signals necessary for the trafficking of the receptor, depends on the quaternary structure of the receptor.
cDNA cloning of a novel cdc2+/CDC28‐related protein kinase from rice
Authors: Hata Shingo
Published: July 2002

A cDNA clone, named R2, has been isolated by screening a rice cell cDNA library with a redundant oligonucleotide probe derived from the conserved ATP binding site of cdc2+/CDC28 protein kinases. The cDNA contained the entire coding sequence for a 424 amino acid polypeptide with a molecular mass of 47.6 kDa. The R2 mRNA, 2.1 kb in size, was expressed in both cultured rice cells and rice seedings at similar levels. The predicted R2 protein has canonical motifs for ATP binding and catalysis, and is significantly homologous (up to 47%) to members of the cdc2+/ CDC28 subfamily of serine/threonine protein kinase. The R2 protein is a novel member of the subfamily.
Carbachol induces secretion in a mast cell line (RBL‐2H3) transfected with the ml muscarinic receptor gene
Authors: Jones S.V.Penelope , Choi Oksoon H. , Beaven Michael A.
Published: July 2002

The possibility that the ml muscarinic receptor subtype can induce release of intracellular granules and transmitters was studied by transfecting a cultured mast cell line, RBL‐2H3 cells, with the ml receptor gene. Comparisons were made between carbachol‐ and antigen‐induced activation of various secretory responses. Like antigen, carbachol stimulated inositol phospholipid hydrolysis and release of arachidonic acid with concomitant dose‐dependent secretion of granular contents. Carbachol also stimulated a biphasic increase in intracellular calcium, as measured by single cell fura‐2 measurements. Although the kinetics of the carbachol‐induced rise in intracellular calcium differed from that induced by antigen, they both utilized the same intracellular pool of calcium, and the second phase of the rise in intracellular calcium was dependent on extracellular calcium in both cases. Thus, the ml muscarinic receptor activates release of granules by a mechanism ostensibly similar to that of antigen.
Identification of 11‐cis and all‐trans‐retinal in the photoreceptive organelle of a flagellate green alga
Authors: Kreimer Georg , Marner Franz-Josef , Brohsonn Uwe , Melkonian Michael
Published: July 2002

Isolation of intact photoreceptive organelles (eyespot apparatuses) involved in blue‐light mediated photoresponses in a flagellate green alga (Spermatozopsis similis) allowed for the first time the identification of both 11‐cis‐ and all‐trans‐retinal in a plant cell. Both isomers were identified by HPLC analysis in conjunction with UV spectra. Additionally, reconstitution of a distinct absorption band, centered around 540 nm, was achieved by addition of exogenous 9‐cis‐retinal to bleached, isolated eyespot apparatuses.
A membrane‐associated cysteine protease inhibitor from murine hepatoma
Authors: Moin Kamiar , Emmert Leslie T. , Sloane Bonnie F.
Published: July 2002

A cysteine protease inhibitor was purified from total membrane fractions of an invasive murine hepatoma, Hepa cl 9. On gel filtration under non‐reducing conditions the purified inhibitor was eluted in a single peak of M1
10–15 kDa, but resolved as two bands at 14 and 70 kDa on SDS‐PAGE under reducing conditions. By isoelectric focusing, the inhibitor ran at an isoelectric point of 4.75. Immunoblotting studies using the enhanced chemiluminescence technique indicated no crossreactivity with monoclonal antibodies to stefin B and cystatin C or with a polyclonal antibody to low M??? kininogen. In contrast, the 14 kDa and 70 kDa bands both crossreacted with a polyclonal antibody to stefin A, suggesting that the cysteine protease inhibitor associated with Hepa cl 9 membranes may be a modified form of stefin A.
Polymerase chain reactions using Saccharomyces, Drosophila and human DNA predict a large family of protein serine/threonine phosphatases
Authors: Chen Mao Xiang , Chen Yu Hua , Cohen Patricia T.W.
Published: July 2002

Using the polymerase chain reaction (PCR) to examine the protein serine/threonine phosphatase (PP) family which includes PP1, PP2A and PP2B, we have identified two, seven, and four novel protein phosphate genes in Saccharomyces cerevisiae, Drosophila melanogaster and Homo sapiens, respectively. Consequently, the genes in the PP1/PP2A/PP2B family now number 11, 15 and 12 in these species respectively, and the data predicts still more unidentified phosphatases in higher eukaryotes. The PCR analyses also point to the presence in Drosophila and mammals of three or more different genes encoding PP2B, the enzyme recently identified as the target of certain immunosuppressant drugs.
Sphingolipid transport from the trans‐Golgi network to the apical surface in permeabilized MDCK cells
Authors: Kobayashi Toshihide , Pimplikar Sanjay W. , Parton Robert G. , Bhakdi Sucharit , Simons Kai
Published: July 2002

We have measured the transport of de novo synthesized fluorescent analogs of sphingomyelin and glucosylceramide from the trans‐Golgi network (TGN) to the apical membrane in basolaterally permeabilized Madin‐Darby canine kidney (MDCK) cells. Sphingolipid transport was temperature, ATP and cytosol dependent. Introduction of bovine serum albumin (BSA), which binds fluorescent sphingolipid monomer, into the permeabilized cells, did not affect lipid transport to the apical membrane. Both fluorescent sphingomyelin and glucosylceramide analogs were localized to the lumenal bilayer leaflet of isolated TGN‐derived vesicles. These results strongly suggest that both sphingolipids are transported from the TGN to the apical membrane via vesicular traffic.
Phenylalanyl‐tRNA synthetase from Thermus thermophilus can attach two molecules of phenylalanine to tRNAPhe
Authors: Stepanov Victor G. , Moor Nina A. , Ankilova Valentina N. , Lavrik Ol'ga I.
Published: July 2002

Phenylalanyl‐tRNA synthetase from the extreme thermophilic bacterium Thermus thermophilus can incorporate more than one molecule of phenylalanine into the tRNAPhe. It is shown that the ‘hyperaminoncylated’ tRNAPPhe is the bis‐2′,3′‐O‐phenylalanyl‐tRNAPhe, and its formation is typical for the thermophilic enzyme but does not occur for E. coli phenylalanyl‐tRNA synthetase under the same conditions.
Structural determination of the active site of a sweet protein A 1H NMR investigation of pMNEI
Authors: Tancredi T. , Iijima H. , Saviano G. , Amodeo P. , Temussi P.A.
Published: July 2002

pMNEI, a single chain sweet protein related to monellin, has been studied by means of 1H NMR at 500 MHz. A partial sequential assignment performed by means of the MCD method allowed the determination of the secondary structure of a large portion of the β‐sheet of pMNEI that contains a likely ‘sweet finger’: the loop connecting the β‐strands from residue 59 to residue 78, corresponding to segment 16–35 of the A chain of monellin. The detailed three‐dimensional structure of the loop (Tyr66‐Ala67‐Ser68‐Asp69), determined from several interresidue and intraresidue NOEs and subsequent energy minimization, shows that the side chains or Tyr66 and Asp69 fit our model of the sweet receptor in a manner very similar to that of the side chains of Phe and Asp or aspartame.
MAP kinase kinase from rabbit skeletal muscle A novel dual specificity enzyme showing homology to yeast protein kinases involved in pheromone‐dependent signal transduction
Authors: Nakielny Sara , Campbell David G. , Cohen Philip
Published: July 2002

MAP kinase kinase (MAPKK) was purified 30,000‐fold to homogeneity from extracts of rabbit skeletal muscle and shown to be a monomeric protein of apparent molecular mass 44 kDa, MAPKK activated the 42 kDa isoform of MAP kinase by phosphorylation of Thr‐183 and Tyr‐185, and phosphorylated itself slowly on tyrosine, threonine and serine residues, establishing that it is a ‘dual specificity’ protein kinase. Peptide sequences from MAPKK were homologous to other protein serine/threonine kinases, especially to the subfamily that includes yeast protein kinase that lie upstream of yeast MAP kinase homologues in the pheromone‐dependent mating pathways.
Nucleolar and cytoplasmic localization of annexin V
Authors: Sun Jiuru , Salem Hatem H. , Bird Philip
Published: July 2002

The subcellular localization of annexin V in cultured human umbilical vein endothelial cells, epithelial cells and fibroblasts was examined. Indirect immunofluorescence and immunoblotting studies using affinity‐purified anti‐annexin V antibodies revealed that annexin V is located within the cytoplasm and nucleus of these cells. Further examination and direct binding studies showed that annexin V within the nucleus is associated with the nucleolus. These findings suggest that annexin V may play a role in a nucleolar function, such as ribosome assembly and transport.
Protegrins: leukocyte antimicrobial peptides that combine features of corticostatic defensins and tachyplesins
Authors: Kokryakov Vladimir N. , Harwig Sylvia S.L. , Panyutich Elena A. , Shevchenko Andrei A. , Aleshina Galina M. , Shamova Olga V. , Korneva Helen A. , Lehrer Robert I.
Published: July 2002

Porcine leukocytes contained three homologous peptides, PG‐1, 2 and 3, that manifested potent microbicidal activity against Escherichia coli, Listeria monocytogenes and Candida albicans in vitro. The peptides (‘protegrins’) were composed of 16 (PG‐2) or 18 amino acid residues, and, like tachyplesins (broad‐spectrum antibiotic peptides of horseshoe crab hemocytes), they contained two intramolecular cystine disulfide bonds. Considerably smaller than defensins, protegrins nevertheless showed substantial homology to them, especially to the ‘corticostatic’ rabbit defensin, NP‐3a. The relatively simple structure of protegrins should provide useful prototypes for constructing congeners with selectively enhanced host defense activities.
The interaction of phthalocyanine with planar lipid bilayers
Authors: Rokitskaya Tatiana I. , Antonenko Yuri N. , Kotova Elena A.
Published: July 2002

The effect of phthalocyanines, the potent photodynamic sensitizers, on the electric properties of the bilayer lipid membrane (BLM) is studied. It is shown, that tetrasulfonated, as well as trisulfonated, aluminium phthalocyanine do not alter the conductance of BLM, but elicit certain changes in the boundary potential difference, which points in favor of dye adsorption on BLM. Under the conditions of intense visible light irradiation, the phthalocyanines cause an increase in the conductance, resulting in the irreversible breakdown of BLM, formed from soy bean phosphati‐dylcholine, but fail to change the conductance of BLM, formed from diphytanoilphosphatidylcholine. The phthalocyanine‐sensitized inactivation of gramicidin channels incorporated into BLM is observed under the conditions of weak visible light irradiation using an He‐Ne laser. The photodynamic blockage of model ionic channels is considerably suppressed after oxygen depletion. The phenomenon consists of a marked reduction of a number of open channels, probably due to photomodification of tryptophan residues, essential for gramicidin functioning. The mechanism of the channel inactivation, involving the photosensitized reaction of the II type, and the relevance to the interaction of sensitizers with biomembranes, is discussed.
The two isoforms of the mouse somatostatin receptor (mSSTR2A and mSSTR2B) differ m coupling efficiency to adenylate cyclase and in agonist‐induced receptor desensitization
Authors: Vanetti Mirko , Vogt Gudrun , Höllt Volker
Published: July 2002

The somatostatin receptor 2 (mSSTR2) is alternatively spliced into two isoforms (mSSTR2A and mSSTR2B) which differ at the C‐terminus. Both receptors bind somatostatin peptides with a similar high affinity when stably expressed in CHO‐K1 cells. However, the spliced form (mSSTR2B) mediates a more efficient inhibition of adenylate cyclase and is much more resistant to agonist‐induced reduction of binding than the longer form (mSSTR2A). These findings indicate that alternative splicing may be a physiological mechanism to modulate receptor desensitization and G‐protein coupling of mSSTR2.
Restricted mobility of the sole tryptophan in membrane‐bound melittin
Authors: Chattopadhyay Amitabha , Rukmini R.
Published: July 2002

In spite of numerous studies, there appears to be no consensus regarding the orientation and aggregation state of membrane‐bound melittin. We report here the restricted environment of the sole tryptophan residue in membrane‐bound melittin using environment‐induced effects on the rates of solvent relaxation. When incorporated into unilamellar vesicles of dioleoyl‐sn‐glycero‐3‐phosphocholine (DOPC), melittin exhibits a red edge excitation shift (REES) of 5 nm. In addition, fluorescence polarization of melittin m the membrane shows both excitation and emission wavelength dependence. Taken together, these observations indicate that the tryptophan residue of melittin is located in a motionally restricted region in the membrane.
Restoration of dystrophin‐associated proteins in skeletal muscle of mdx mice transgenic for dystrophin gene
Authors: Matsumura Kiichiro , Lee Cheng Chi , Caskey C.Thomas , Campbell Kevin P.
Published: July 2002

Duchenne muscular dystrophy (DMD) patients and mdx mice are characterized by the absence of dystrophin, a membrane cytoskeletal protein. Dystrophin is associated with a large oligomeric complex of sarcolemmal glycoproteins, including dystroglycan which provides a linkage to the extarcellular matrix component, laminin. The finding that all of the dystrophin‐associated proteins (DAPs) are drastically reduced in DMD and mdx skeletal muscle supports the primary function of dystrophin as an anchor of the sarcolemmal glycoprotein complex to the subsarcolemmal cytoskeleton. These findings indicate that the efficacy of dystrophin gene therapy will depend not only on replacing dystrophin but also on restoring all of the DAPs in the sarcolemma. Here we have investigated the status of the DAPs in the skeletal muscle of mdx mice transgenic for the dystrophin gene. Our results demonstrate that transfer of dystrophin gene restores all of the DAPs together with dystrophin, suggesting that dystrophin gene therapy should be effective in restoring the entire dystrophin‐glycoprotein complex.
Extended in vivo blood circulation time of fluorinated liposomes
Authors: Santaella Catherine , Frézard Frédéric , Vierling Pierre , Riess Jean G.
Published: July 2002

The clearance from blood circulation of fluorinated liposomes made with perfluoroalkylated phosphatidylcholines was investigated in mice using liposome‐entrapped 5(6)‐carboxyfluorescein. The presence of a fluorinated core inside the membrane strongly retards their blood clearance. The fluorinated vesicles showed circulation half‐lives of up to 8.6 h, which are 6–13 and 3–6 times larger than those of similarly sized conventional distearoylphosphatidylcholine and distearoylphosphatidylcholine/cholesterol liposomes, respectively. Their blood clearance was similar to that of some polyethylene glycol (PEG)‐labelled ‘stealth’ liposomes and was dose‐independent in a 3.3–330 body weight dose range.
The lysosomal gp63‐related protein in Leishmania mexicana amastigotes is a soluble metalloproteinase with an acidic pH optimum
Authors: Ilg Thomas , Harbecke Dorothee , Overath Peter
Published: July 2002

Leishmania mexicana amastigotes express a lysosomal protein, which is antigenically related to the promastigote surface metalloproteinase (gp63). It is shown that the purified gp63‐related protein from amastigote is also an active metalloproteinase. The pH‐optimum of the enzyme is acidic, similar to lysosomal cysteine proteinases, but distinct from the neutral to basic pH‐optimum of the promastigote surface proteinase. This study appears to be the first report on a metalloproteinase with a lysosomal localization.
The Rhodobacter sphaeroides PufX protein is not required for photosynthetic competence in the absence of a light harvesting system
Authors: McGlynn Peter , Hunter C.Neil , Jones Michael R.
Published: July 2002

The effects of deletion of the gene encoding the PufX protein from Rhodobacter sphaeroides have been examined using bacterial strains with simplified photosystems. We find that the PufX protein is required for photosynthetic growth in strains which have the LH1 antenna complex, but is not required in a reaction centre‐only strain, suggesting that the PufX protein does not directly facilitate cyclic electron transfer between the reaction centre and the cytochrome bc
1 complex. The influence of PufX and carotenoid type on the size of the reaction center/LH1 core complex has also been examined in these strains.
A possible role of sphingosine in induction of apoptosis by tumor necrosis factor‐α, in human neutrophils
Authors: Ohta Hideki , Yatomi Yutaka , Sweeney Elizabeth A. , Hakomori Sen-itiroh , Igarashi Yasuyuki
Published: July 2002

Treatment of human neutrophils with tumor necrosis factor‐α (TNF‐α) resulted in an increase in concentration of ceramide and its catabolite, sphingosine. Sphingosine, a potent endogenous protein kinase C (PKC) inhibitor, as well as TNF‐α, induced internucleosomal DNA fragmentation and morphological changes characteristic of apoptotic cells. Ceramide and sphingosine‐1‐phosphate, the initial product of sphingosine catabolism, did not cause apoptosis under our experimental conditions. In addition, 1‐(5‐isoquinolinesulfonyl)‐2‐methylpiperazine (H7) and N,N‐dimethylsphingosine (DMS), known as PKC inhibitors, also induced apoptosis, suggesting that induction of apoptosis by sphingosine may be related to inhibition of PKC activity. These results indicate that sphingosine deacylated from ceramide may be an endogenous modulator mediating apoptotic signals by TNF‐α in neutrophils.
Purification, inhibitory properties, amino acid sequence and identification of the reactive site of a new serine proteinase inhibitor from oil‐rape (itBrassica napus) seed
Authors: Ceciliani Fabrizio , Bortolotti Fabrizio , Menegatti Enea , Ronchi Severino , Ascenzi Paolo , Palmieri Sandro
Published: July 2002

A new serine proteinase inhibitor, rapeseed trypsin inhibitor (RTI), has been isolated from rapeseed (Brassica napus var. oleifera) seed. The protein inhibits the catalytic activity of bovine β‐trypsin and bovine α‐chymotrypsin with apparent dissociation constants of 3.0 × 10−10 M and 4.1 × 10−7 M, at pH 8.0 and 21°C, respectively. The stoichiometry of both proteinase‐inhibitor complexes is 1:1. The amino acid sequence of RTI consists of 60 amino acid residues, corresponding to an M
r, of about 6.7 kDa. The p1‐pi, reactive site bond has been tentatively identified at position Arg20‐Ile21. RTI shows no similarity to other serine proteinase inhibitors except the low molecular weight mustard trypsin inhibitor (MTI‐2). RTI and MTI‐2 could be members of a new class of plant serine proteinase inhibitors.
Evidence for trans‐cis isomerization of the p‐coumaric acid chromophore as the photochemical basis of the photocycle of photoactive yellow protein
Authors: Kort R , Vonk H , Xu X , Hoff W.D , Crielaard W , Hellingwerf K.J
Published: July 2002

Analysis of the chromophore p‐courmaix acid, extracted from the ground state and the long‐lived blue‐shifted photocycle intermediate of photoactive yellow protein, shows that the chromophore is reversibly converted from the trans to the cis configuration, while progressing through the photocycle. The detection of the trans and cis isomers was carried out by high performance capillary zone electrophoresis and further substained by 1H NMR spectroscopy. The data presented here establish the photo‐isomerization of the vinyl double bond in the chromophore yellow protein, a eubacterial photosensory protein. A similar isomerization process occurs in the structurally very different sensory rhodopsins, offering an explanation for the strong spectroscopic similarities between photoactive yellow protein and the sensory rhodopsins. This is the first demonstration of light‐induced isomerization of a chromophore double bond as the photochemical basis for photosensing in the domain of Bacteria.
Initial analysis of 750 MHz NMR spectra of selectively 15N‐G,U labelled E. coli 5S rRNA
Authors: Grüne Matthias , Görlach Matthias , Soskic Vukic , Klussmann Sven , Bald Rolf , Fürste Jens Peter , Erdmann Volker A. , Brown Larry R.
Published: July 2002

The overall folding of an RNA molecule is reflected in its base pairing pattern. The identification of that pattern provides a first step towards the determination of the structure of an RNA molecule. We show that the application of heteronuclear NMR methods at 750 MHz to E. coli 5S rRNA (120 nucleotides) selectively labelled with 15N in guanine and uridine allows observation of base pairing patterns for a larger RNA molecule. We also present evidence that the fold of the E‐domain of the 5S rRNA (nt 79–97) as a contiguous part of the 5S rRNA and as an isolated molecule is virtually the same.
Multiple inhibitory effects of Alzheimer's peptide Aβ1–40 on lipid biosynthesis in cultured human HepG2 cells
Authors: Koudinova Natalia V , Berezov Temirbolat T , Koudinov Alexei R
Published: July 2002

Herein we describe the inhibitory effect of the synthetic peptide Aβ1–40, homologous to the major high‐density lipoprotein‐associated species of Alzheimer's amyloid β protein (Aβ), on lipid biosynthesis in human hepatic HepG2 cells. This culture synthesizes various lipids from [14C]acetate as a precursor. Treatment of cells with different concentrations of Aβ1–40 decreased the syntheses of various radiolabeled lipid species. The decrease reached saturation at peptide concentrations equal to 10–100 ng ml−1. The lipids whose synthesis was decreased most were free and esterified cholesterol and phospholipids. This inhibitory effect suggests that Aβ protein may modulate physiological intracellular lipid syntheses. It may also be of special importance in the pathological condition, and contribute to the neurodegeneration, in Alzheimer's disease and related disorders.
ATPase cycle controls the conformation of an archaeal chaperonin as visualized by cryo‐electron microscopy
Authors: Gutsche Irina , Mihalache Oana , Hegerl Reiner , Typke Dieter , Baumeister Wolfgang
Published: July 2002

Chaperonins are double‐ring protein folding machines fueled by ATP binding and hydrolysis. Conformational rearrangements upon ATPase cycling of the group I chaperonins, typified by the Escherichia coli GroEL/GroES system, have been thoroughly investigated by cryo‐electron microscopy and X‐ray crystallography. For archaeal group II chaperonins, however, these methods have so far failed to provide a correlation between the structural and the functional states. Here, we show that the conformation of the native αβ‐thermosome of Thermoplasma acidophilum in vitrified ice is strictly regulated by adenine nucleotides.
Chloride dependence of glycine betaine transport in Halobacillus halophilus
Authors: Roeßler Markus , Müller Volker
Published: July 2002

Growth of Halobacillus halophilus is strictly chloride‐dependent but the physiological basis for the chloride dependence remains to be elucidated. To address the function of Cl− in H. halophilus, a physiological study was performed. It was found that uptake of the compatible solute glycine betaine under isoosmotic conditions was stimulated by increasing salt concentrations. Uptake of glycine betaine required both, Na+ and Cl−. Cl− could be substituted by nitrate and bromide, but not by sulfate. Glycine betaine transport was optimal at around 0.7 M Cl−. Cells responded to an osmotic upshock by accumulating glycine betaine, but only in the presence of chloride. These studies revealed the first chloride‐dependent glycine betaine transporter in a prokaryote.
An estimation of the size of the water cluster present at the cleavage site of the water splitting enzyme
Authors: Burda Kvetoslava , Bader Klaus P. , Schmid Georg H.
Published: July 2002

In time‐dependent measurements of oxygen evolution in tobacco thylakoid membranes we varied the fraction of H2
18O and the temperature and measured water splitting as 18O2, 16O18O, and 16O2 by mass spectrometry. We show that the approach to the equilibrium of the system after H2
18O addition can be very well understood in terms of the diffusion of water molecules. The equilibrium states of 16O2, 16O18O, and 18O2 evolution differ from the theoretical binomial distributions, which are expected under the prerequisite of ideal mixing of the water molecules and that of the chemical equivalence of H2
18O and H2
16O for an infinite cluster. The presence of this deviation means that there is a typical size of water clusters having access to cleavage by the water splitting enzyme. We estimated that this cluster contains about 12±2 water molecules.
Nanoscale biogenic iron oxides and neurodegenerative disease
Authors: Dobson Jon
Published: July 2002

One of the characteristics of many neurodegenerative diseases is the disruption of normal iron homeostasis in the brain. Recent experimental work indicates that nanoscale magnetic biominerals (primarily magnetite and maghemite) may be associated with senile plaques and tau filaments found in brain tissue affected by these diseases. These findings have important implications for our understanding of the role of iron in neurodegenerative disease as well as profound implications for their causes. In addition, the presence of biogenic magnetite in affected tissue should also provide improved mechanisms for early detection through the modification of MRI pulse sequences.
Mechanical unfolding of single filamin A (ABP‐280) molecules detected by atomic force microscopy
Authors: Furuike Shou , Ito Tadanao , Yamazaki Masahito
Published: July 2002

Filamin A (ABP‐280), which is an actin‐binding protein of 560 kDa as a dimer, can, together with actin filaments, produce an isotropic cross‐linked three‐dimensional network (actin/filamin A gel) that plays an important role in mechanical responses of cells in processes such as maintenance of membrane stability and translational locomotion. In this study, we investigated the mechanical properties of single filamin A molecules using atomic force microscopy. In force–extension curves, we observed sawtooth patterns corresponding to the unfolding of individual immunoglobulin (Ig)‐fold domains of filamin A. At a pulling speed of 0.37 μm/s, the unfolding interval was sharply distributed around 30 nm, while the unfolding force ranged from 50 to 220 pN. This wide distribution of the unfolding force can be explained by variation in values of activation energy and the width of activation barrier of 24 Ig‐fold domains of the filamin A at the unfolding transition. This unfolding can endow filamin A with great extensibility. The refolding of the unfolded chain of filamin A occurred when the force applied to the protein was reduced to near zero, indicating that its unfolding is reversible. Based on these results, we discuss here the physiological implications of the mechanical properties of single filamin A molecules.
Molecular cloning of ERp29, a novel and widely expressed resident of the endoplasmic reticulum
Authors: Demmer Jerome , Zhou ChaoMing , Hubbard Michael J
Published: July 2002

We have isolated a full‐length cDNA clone for a novel 29 kDa protein that is highly expressed in rat enamel cells. The clone encodes a 259‐residue protein, here named ERp29, with structural features (signal peptide and a variant endoplasmic reticulum‐retention motif, KEEL) that indicate it is a reticuloplasmin. ERp29 has limited homology with protein disulfide isomerase and its cognates, but lacks their characteristic thioredoxin‐like catalytic moiety and calcium‐binding motifs. ERp29 mRNA was expressed in all rat tissues tested, and a homologous transcript was detected in other animal livers (primate, ruminant, marsupial). In human hepatoma cells, ERp29 mRNA expression was not increased by stresses (tunicamycin, calcium ionophore) that induced other reticuloplasmins. We conclude that ERp29 is a new, highly conserved member of the reticuloplasmin family which is widely expressed. The apparent lack of both calcium binding properties and stress responsiveness distinguish ERp29 from all major reticuloplasmins characterised to date.
Differential expression of tissue transglutaminase during in vivo apoptosis of thymocytes induced via distinct signalling pathways
Authors: Szondy Zsuzsa , Molnar Peter , Nemes Zoltán , Boyiadzis Michael , Kedei Noémi , Tóth Réka , Fésüs László
Published: July 2002

© 1997 Federation of European Biochemical Societies.
A novel and ancient vertebrate opsin
Authors: Soni Bobby G , Foster Russell G
Published: July 2002

We describe the identification of a novel opsin gene isolated from the eyes of Atlantic salmon. The cDNA sequence predicts a protein that has the key features of an opsin, but shows only 32–42% amino acid identity to the known opsin families. Phylogenetic analysis suggests that this opsin is a member of a hitherto unrecognised opsin family that diverged early in the evolution of vertebrate photopigments. We have tentatively called this opsin family the vertebrate ancient (VA) opsins. The identification of VA opsin may ultimately help to resolve some of the uncharacterised photoreceptor functions of the eye, which include the regulation of circadian rhythms, pupil size and corneal pigmentation.
Crystallization, preliminary diffraction and electron paramagnetic resonance studies of a single crystal of cytochrome P450nor
Authors: Park Sam-Yong , Shimizu Hideaki , Adachi Shin-ichi , Yoshitsugu Shiro  , Iizuka Tetsutaro , Nakagawa Atsushi , Tanaka Isao , Shoun Hirofumi , Hori Hiroshi
Published: July 2002

Cytochrome P450nor (P450nor) is a heme‐containing nitric oxide reductase from the denitrifying fungus, Fusarium oxysporum. This enzyme catalyzes the reduction of NO to N2O. In the present study, we report results from preliminary crystallographic and electron paramagnetic resonance (EPR) analysis of a single crystal of P450nor. The crystal was grown in 100 mM MES buffer at pH 5.6 using PEG 4000 as a precipitant. It belongs to the orthorhombic system with cell dimensions of a=54.99 Å, b=82.66 Å, c=87.21 Å, and the space group is P212121. The crystal diffracts synchrotron radiation at higher than 2.0 Å resolution, and therefore it is suitable for X‐ray crystal structure analysis at atomic resolution. Bijvoet and dispersive anomalous difference Patterson maps show a clear peak corresponding to the heme iron. The structure solution is currently underway by means of MIR and MAD techniques. EPR analysis determined the orientation of the heme within the P450nor crystal.
Glutamine accelerates interleukin‐6 production by rat peritoneal macrophages in culture
Authors: Yassad Afef , Lavoinne Alain , Bion Anne , Daveau Maryvonne , Husson Annie
Published: July 2002

The effect of glutamine on the production of interleukin‐6 (IL‐6) was studied in rat peritoneal macrophages in culture. A maximal production of IL‐6 was measured at 4 h in lipopolysaccharide (LPS)‐stimulated macrophages, and addition of glutamine (5 mM) anticipated this increase by 1 h without any increase in the IL‐6 mRNA level. The effect of glutamine required the presence of LPS. Thus, glutamine accelerates IL‐6 production from the pre‐existing mRNA. The effect of glutamine was not mediated by cell swelling since culture of macrophages in hypoosmotic condition decreased the production of IL‐6 in the culture medium with a corresponding decrease in the IL‐6 mRNA level.
Differential internalisation of mGluR1 splice variants in response to agonist and phorbol esters in permanently transfected BHK cells
Authors: Ciruela F. , McIlhinney R.A.J.
Published: July 2002

The internalisation of metabotropic glutamate receptor (mGluR1α) and its splice variant (mGluR1β), in response to agonist and phorbol esters (PMA), has been studied. Both mGluR1α and mGluR1β exhibit a similar rate of internalisation following PMA treatment, with a shift in their distribution from plasma membrane to endosome‐enriched membrane fractions. Agonist challenge however caused a rapid loss, within 5–10 min, of mGluR1β but not mGluR1α from the cell surface. These results show that the two forms of mGluR1 show different internalisation responses to agonist and suggest that the C‐terminal region of the molecule plays an important role in this phenomenon.
Functional expression of pharaonis phoborhodopsin in Eschericha coli
Authors: Shimono Kazumi , Iwamoto Masayuki , Sumi Masato , Kamo Naoki
Published: July 2002

Pharaonis phoborhodopsin, the photoreceptor of the negative phototaxis of archaebacterial Natronobacterium pharaonis, was functionally expressed in the heterologous system of Escherichia coli. Flash‐photolysis on a millisecond time scale indicated that the photochemical properties of ppR expressed in E. coli were the same as those of the native ppR in N. pharaonis. We concluded that the integral membrane protein ppR is correctly folded in vivo in the eubacterial E. coli membrane.
Anti‐peptide antibodies specific to rat endothelin‐converting enzyme‐1 isoforms reveal isoform localisation and expression
Authors: Brown Carolyn D , Barnes Kay , J. Turner Anthony
Published: July 2002

Endothelin‐converting enzyme‐1 (ECE‐1) is a critical enzyme in the biosynthesis of the potent vasoconstrictor peptide endothelin and exists in several isoforms. Anti‐peptide antibodies raised against epitopes in the distinct N‐terminal cytoplasmic tails of the rat ECE‐1 isoforms have been obtained. By using these antibodies in Western blot analysis and immunofluorescence studies, we have shown that cultures of transformed rat lung vascular endothelial cells treated with the metalloprotease inhibitor phosphoramidon and untreated cells express ECE‐1α only, whereas human umbilical vein endothelial cells express ECE‐1α and ECE‐1β. The ECE‐1 isoforms expressed in CHO‐K1 cells transfected with rat cDNA to ECE‐1α and ECE‐1β could be immunoprecipitated by using the appropriate isospecific antibody.