bluuebunny/crossref_metadata_2025 huggingface dataset:
Cytosolic factors mediate protein insertion into the peroxisomal membrane
Authors: Pause Birgit , Diestelkötter Petra , Heid Hans , Just Wilhelm W.
Published: July 2002

Following in vitro translation of the 22 kDa peroxisomal membrane protein (Pmp22p), gel filtration analysis of the post‐ribosomal supernatant revealed that Pmp22p forms two complexes. Complex I is of high molecular weight, results in a crosslinking product of 80 kDa, and by co‐immunoprecipitation with anti‐TCP1 antibody was identified as TRiC. In complex II Pmp22p was crosslinked to a yet unknown polypeptide of 40 kDa (P 40). This complex exhibited much higher efficiency to insert Pmp22p into the peroxisomal membrane compared to complex I. In a model we suggest that newly synthesized Pmp22p is first bound to TRiC before being transferred to P 40 which may function as a cytosolic Pmp22p receptor.
The tomato RBCS3A promoter requires integration into the chromatin for correct organ‐specific regulation
Authors: Meier Iris , Gröning Birge , Michalowski Susan , Spiker Steven
Published: July 2002

In tomato, the RBCS1, RBCS2 and RBCS3A genes, encoding the small subunit of ribulose‐1,5‐bisphosphate carboxylase/oxygenase, are expressed in leaves and light‐grown seedlings, but only RBCS1 and RBCS2 are expressed in developing tomato fruits. The activities of the three promoters have been compared in transgenic plants and after transient transformation. Fruit‐specific repression of the RBCS3A promoter was observed in transgenic plants, but not after ballistic transient transformation, indicating that chromatin integration is necessary for its correct organ‐specific regulation. In addition, matrix attachment regions have been identified in the RBCS1, RBCS2 and RBCS3A promoters. This is the second case in plants of absence of correct regulation of a plasmid‐borne plant promoter and correlating potential nuclear matrix attachment of the gene.
The disulphide bond pattern of bitistatin, a disintegrin isolated from the venom of the viperBitis arietans
Authors: Calvete Juan J , Schrader Michael , Raida Manfred , McLane Mary Ann , Romero Antonio , Niewiarowski Stefan
Published: July 2002

The disulphide bond pattern of the long disintegrin bitistatin (83 amino acids, 14 cysteines) was established using structural information gathered by amino acid analysis, N‐terminal sequencing, and molecular mass determination of fragments isolated by reversed‐phase HPLC after polypeptide degradation with trypsin and oxalic acid. A computer program was used to calculate all possible combinations of disulphide‐bonded peptides matching the mass spectrometric data, and the output was filtered using compositional and sequence data. Disulphide bonds between cysteines 16–34, 18–29, 28–51, 42–48, 47–72, and 60–79 are conserved in medium‐long disintegrins flavoridin and kistrin (70 amino acids, 12 cysteines), and the two cysteine residues at positions 5 and 24 found in bitistatin but not in other disintegrin molecules are disulphide‐bridged. This linkage creates an extra, large loop, which, depending on whether the NMR structure of flavoridin or kistrin is used for modelling the structure of bitistatin, lies opposite or nearly parallel, respectively, to the biologically active RGD‐containing loop.
Hemin and related porphyrins inhibit β‐amyloid aggregation
Authors: Howlett David , Cutler Paul , Heales Simon , Camilleri Patrick
Published: July 2002

Porphyrins related to the naturally occurring pigment heme were found to effectively interfere with the aggregation of β‐amyloid peptides as determined by an immunoassay configured for the detection of β‐amyloid oligomers. Oligomerisation of β‐amyloid is believed to be a key event in the progression of Alzheimer's disease. Inhibition of this aggregation is thus an important strategy in combating this commonest form of senile dementia. Evidence was also generated for hemin and hematin mediated protection of cultured cells against the neurotoxic effects of β‐amyloid. These data are discussed with reference to the known pathology of Alzheimer's disease and the chemistry of porphyrins.
A novel Bacillus natto plasmid pLS32 capable of replication in Bacillus subtilis
Authors: Tanaka Teruo , Ogura Mitsuo
Published: July 2002

Plasmid pLS32 is a relatively large (approximately 70 kbp), cryptic, low copy‐number plasmid present in Bacillus natto. We isolated and analyzed the replication region of the plasmid in B. subtilis, and the following results were obtained: the replication region contained an open reading frame encoding a 287‐amino acid protein (RepN), whose amino acid sequence was partially homologous with those of the Rep proteins encoded on plasmids pAD1 and pLJ1 isolated from Enterococcus faecalis and Lactobacillus helveticus, respectively; the replication origin (oriN) was located in the repN‐coding region; the copy number of a pLS32 derivative, pBET131, was 2 to 3 per chromosome; replication of pBET131 required polI. These features show that pLS32 is a novel plasmid capable of replication in B. subtilis.
Vitronectin‐dependent invasion of epithelial cells by Neisseria gonorrhoeae involves αv integrin receptors
Authors: Dehio Michaela , Gómez-Duarte Oscar G , Dehio Christoph , Meyer Thomas F
Published: July 2002

Binding of vitronectin (VN) to Neisseria gonorrhoeae expressing the heparan sulfate proteoglycan (HSPG) specific Opa50 protein was recently shown to trigger bacterial internalization into distinct epithelial cell lines. We have investigated the role of VN‐binding integrin receptors and protein kinase C (PKC) in VN‐triggered bacterial uptake. Blocking integrin function by RGDS peptides or by antibodies specific to αvβ5 or αvβ3 resulted in an abrogation of VN‐triggered bacterial internalization. Moreover, inhibitors of PKC were found to block VN‐triggered uptake. The essential role of αv integrins and the presumable involvement of PKC in VN‐triggered gonococcal uptake are discussed.
G‐protein βγ‐binding domains regulate insulin exocytosis in clonal pancreatic β‐cells
Authors: Zhang Hui , Yasrebi-Nejad Houri , Lang Jochen
Published: July 2002

We have tested the putative role of G‐protein β‐subunits in insulin exocytosis by transient expression of βγ‐binding proteins targeted to the plasma membrane. The PH domain of the G‐protein‐linked receptor kinase 2 fused to the transmembrane domain of a cell surface receptor and the α‐subunit of the retinal G‐protein transducin inhibited stimulated insulin release from intact and permeabilised HIT‐T15 cells. This effect cannot be imputed to an increase in free Gα, as the RGS protein RGS3 did not reverse this effect. Among the isoforms of Gβ examined, Gβ2 was detected on the plasma membrane by confocal immunomicroscopy. These observations suggest a role for G‐protein βγ‐subunits in insulin exocytosis.
Polypyrimidine tract‐binding protein interacts with HnRNP L
Authors: Hahm Bumsuk , Cho Ook H , Kim Jung-E , Kim Yoon K , Kim Jong H , Oh Young L , Jang Sung K
Published: July 2002

Polypyrimidine tract‐binding protein (PTB) is involved in pre‐mRNA splicing and internal ribosomal entry site (IRES)‐dependent translation. In order to identify cellular protein(s) interacting with PTB, we performed a yeast two‐hybrid screening. Heterogeneous nuclear ribonucleoprotein L (hnRNP L) was identified as a PTB‐binding protein. The interaction between PTB and hnRNP L was confirmed in an in vitro binding assay. Both PTB and hnRNP L were found to localize in the nucleoplasm, excepting the nucleoli, in HeLa cells by the green fluorescent protein (GFP)‐fused protein detection method. The N‐terminal half of PTB (aa 1–329) and most of hnRNP L (aa 141–558) is required for the interaction between PTB and hnRNP L.
Cytochrome c release and caspase activation in hydrogen peroxide‐ and tributyltin‐induced apoptosis
Authors: Stridh Hélène , Kimland Monica , Jones Dean P. , Orrenius Sten , Hampton Mark B.
Published: July 2002

The ability of H2O2 and tributyltin (TBT) to trigger pro‐caspase activation via export of cytochrome c from mitochondria to the cytoplasm was investigated. Treatment of Jurkat T lymphocytes with H2O2 resulted in the appearance of cytochrome c in the cytosol within 2 h. This was at least 1 h before caspase activation was observed. TBT caused cytochrome c release already after 5 min, followed by caspase activation within 1 h. Measurement of mitochondrial membrane potential (ΔΨm) showed that both H2O2 and TBT dissipated ΔΨm, but with different time courses. TBT caused a concomitant loss of ΔΨm and release of cytochrome c, whereas cytochrome c release and caspase activation preceded any apparent ΔΨm loss in H2O2‐treated cells. Thus, our results suggest that different mechanisms are involved in triggering cytochrome c release with these apoptosis‐inducing agents.
Differential actin organization by vinculin isoforms: implications for cell type‐specific microfilament anchorage
Authors: Rüdiger Manfred , Korneeva Nadja , Schwienbacher Christine , Weiss Elisabeth E , Jockusch Brigitte M
Published: July 2002

Vinculin is found in all adherens junctions, while metavinculin, a larger splice variant, is coexpressed with vinculin only in smooth and cardiac muscle. To understand the significance of metavinculin expression, we compared ligand binding between turkey vinculin and metavinculin. Residues 1–258 were found essential for head‐tail interactions in both proteins. The tail domains (VT and MVT, respectively) both bind to F‐actin. However, while VT bundles F‐actin, MVT generates highly viscous F‐actin webs. In transfected PtK2 cells, VT causes F‐actin needles or coils, while MVT‐expressing cells display a diffuse F‐actin distribution. Thus, the MVT‐specific insert induces an F‐actin supraorganization different from the VT‐based form, suggesting that metavinculin has a specific role in muscle.
Nitric oxide induces and inhibits apoptosis through different pathways
Authors: Shen Ying H , Wang Xing L , Wilcken David E.L
Published: July 2002

Physiological levels of nitric oxide (NO) regulate vascular tone and protect the microvasculature from injury whereas excessive NO may be harmful. The present study explored the effects of NO on human endothelial cell apoptosis. We found that the NO donor S‐nitroso‐N‐acetylpenicillamine (SNAP) inhibited TNFα‐induced endothelial apoptosis and that this was mediated partly through the cGMP pathway. In contrast, high SNAP concentration induced endothelial apoptosis via cGMP‐independent pathways and the cGMP pathway protected against NO‐induced apoptosis. These findings demonstrate that low NO concentrations contribute to human endothelial cell survival, whereas higher NO concentrations are pathological and promote destruction of endothelial cells.
Inhibition of Ca2+‐independent phospholipase A2 by bromoenol lactone attenuates prostaglandin generation induced by interleukin‐1β and dibutyryl cAMP in rat mesangial cells
Authors: Akiba Satoshi , Hayama Misako , Sato Takashi
Published: July 2002

Cytokine‐induced prostaglandin generation in rat mesangial cells has been suggested to be dependent on the expression of secretory phospholipase A2 (sPLA2). In the present study, we investigated the possible involvement of Ca2+‐independent phospholipase A2 (iPLA2) in the generation. The results showed that bromoenol lactone, a relatively selective iPLA2 inhibitor, significantly attenuated prostaglandin E2 generation induced by interleukin‐1β and dibutyryl cAMP in parallel with the inhibition of iPLA2 activity. However, the inhibitor did not affect sPLA2 release upon stimulation, activities of sPLA2 or cytosolic phospholipase A2, or Ca2+ ionophore‐induced arachidonic acid liberation. These results suggest that prostaglandin E2 generation upon stimulation may be partially mediated by iPLA2 in addition to sPLA2.
Isolation and characterization of circulating 13‐kDa C‐terminal fragments of human insulin‐like growth factor binding protein‐5
Authors: Ständker Ludger , Wobst Philip , Mark Silke , Forssmann Wolf-Georg
Published: July 2002

The insulin‐like growth factor binding proteins (IGFBPs) are responsible for regulation of the effects and the bioavailability of the insulin‐like growth factors (IGFs). We screened for circulating fragments of human IGFBP‐5 in human hemofiltrate. Identification of IGFBP‐5 peptides in the fractions of our peptide bank generated from hemofiltrate was performed by their immunoreactivity and their capacity to bind IGF‐I. Different fragments of IGFBP‐5 with molecular sizes from 12 to 25 kDa were identified. C‐terminal peptides of IGFBP‐5 with molecular masses of 13.3 and 13.5 kDa were purified by consecutive chromatographic steps and sequenced. Sequence analysis of the peptides revealed the (double) sequences (K)FVGGAENXAHPRII and MVPRAVYLPNXDRKG. In addition, a smaller fragment with M
r 2722 of the central IGFBP‐5 region was purified and showed the sequence HTRISELKAEAVKKDRRKKLTQS (residues 121–143) indicating plasma proteolysis of IGFBP‐5 C‐terminal to amino acids Lys‐120, Ser‐143, Lys‐144, and Arg‐188. According to mass spectrometric and sequence analysis, Thr‐152 was shown to be O‐glycosylated. Fractions containing C‐terminal IGFBP‐5 fragments revealed significant IGF‐I binding properties. Our results indicate that plasma proteolysis of IGFBP‐5 preferentially occurs C‐terminally to basic residues and generates different C‐terminal fragments, possibly acting in an IGF‐dependent manner and bearing intrinsic biological functions.
Transition protein 4 from boar late spermatid nuclei is a topological factor that stimulates DNA‐relaxing activity of topoisomerase I
Authors: Akama Kuniko , Kondo Miyuki , Sato Hiroki , Nakano Minoru
Published: July 2002

Transition protein 4 (TP4) from boar late spermatid nuclei, having higher affinity for double‐stranded DNA and a local melting activity of DNA, stimulated SV40 DNA‐relaxing activity of eukaryotic topoisomerase I at TP4/DNA molar ratios of 6.6–11. A TP4‐spermidine mixture stimulated the activity of topoisomerase I much more than spermidine alone, but no more than TP4 alone, and poly‐l‐arginine did not. These results suggest that TP4 contributes to the chromatin reorganization in the late spermatid nuclei from nucleosomal‐type structure with negatively supercoiled DNA to nucleoprotamine structure with no supercoiled DNA.
Stimulation of the catalytic activity of poly(ADP‐ribosyl) transferase by transcription factor Yin Yang 1
Authors: Griesenbeck Joachim , Ziegler Mathias , Tomilin Nikolai , Schweiger Manfred , Oei Shiao Li
Published: July 2002

The transcriptional regulator Yin Yang 1 (YY1) has previously been demonstrated to physically interact with poly(ADP‐ribosyl) transferase (ADPRT). This nuclear enzyme catalyzes the synthesis of ADP‐ribose polymers and their attachment to target proteins. It is reported here that YY1 associates preferably with the extensively auto(ADP‐ribosyl)ated form of ADPRT, but not with deproteinized ADP‐ribose polymers. In the presence of YY1 the catalytic rate of ADPRT is enhanced about 10‐fold. This stimulation is in part due to modification of YY1, thus serving as a substrate of the reaction. In addition, automodification of ADPRT is also substantially increased. The activation by YY1 is most pronounced at low concentrations of ADPRT suggesting that the presence of YY1 may either facilitate the formation of catalytically active dimers of ADPRT or lead to the occurrence of active heterooligomers. The potential significance of these observations was verified by analyzing the activity of ADPRT in HeLa nuclear extracts. The endogenous enzyme exhibited an about 10‐fold higher activity as compared to the isolated recombinant protein. It is likely that the heat‐stable transcription factor YY1 contributed to the increased activity of ADPRT detected in the nuclear extracts, because heated extracts had a similar stimulatory effect on isolated ADPRT as isolated YY1 used at comparable concentrations. It is concluded that YY1 may be an important regulator of ADPRT and, therefore, could support the function of ADPRT to facilitate DNA repair.
Phosphorylation of yeast TBP by protein kinase CK2 reduces its specific binding to DNA
Authors: Maldonado Edio , Allende Jorge E
Published: July 2002

Protein kinase CK2 is a ubiquitous Ser/Thr kinase which phosphorylates a large number of proteins including several transcription factors. Recombinant Xenopus laevis CK2 phosphorylates both recombinant Saccharomyces cerevisiae and Schizosaccharomyces pombe TATA binding protein (TBP). The phosphorylation of TBP by CK2 reduces its binding activity to the TATA box. CK2 copurifies with the transcription factor IID (TFIID) complex from HeLa cell extracts and phosphorylates several of the TBP‐associated factors within TFIID. Taken together these findings argue for a role of CK2 in the control of transcription by RNA polymerase II through the modulation of the binding activity of TBP to the TATA box.
The Enterococcus hirae copper chaperone CopZ delivers copper(I) to the CopY repressor
Authors: Cobine Paul , Wickramasinghe Wasantha A. , Harrison Mark D. , Weber Thomas , Solioz Marc , Dameron Charles T.
Published: July 2002

Expression of the cop operon which effects copper homeostasis in Enterococcus hirae is controlled by the copper responsive repressor CopY. Purified Zn(II)CopY binds to a synthetic cop promoter fragment in vitro. Here we show that the 8 kDa protein CopZ acts as a copper chaperone by specifically delivering copper(I) to Zn(II)CopY and releasing CopY from the DNA. As shown by gel filtration and luminescence spectroscopy, two copper(I) are thereby quantitatively transferred from Cu(I)CopZ to Zn(II)CopY, with displacement of the zinc(II) and transfer of copper from a non‐luminescent, exposed, binding site in CopZ to a luminescent, solvent shielded, binding site in CopY.
A single uridine modification at the wobble position of an artificial tRNA enhances wobbling in an Escherichia coli cell‐free translation system
Authors: Takai Kazuyuki , Okumura Shuhei , Hosono Kazumi , Yokoyama Shigeyuki , Takaku Hiroshi
Published: July 2002

5‐Methoxyuridine was introduced into the first position of the anticodon of the unmodified form of tRNA1
Ser from Escherichia coli. The codon reading efficiencies of this tRNA (tRNA(5‐methoxyuridine UGA)) relative to those of the unmodified counterpart (tRNA(UGA)) were measured in a cell‐free translation system. tRNA(5‐methoxyuridine UGA) was more efficient than tRNA(UGA) in the reading of the UCU and UCG codons and was less efficient in the reading of the UCA codon. Thus, the single modification of U to 5‐methoxyuridine can enhance the wobble readings.
Polyunsaturated fatty acids potentiate interleukin‐1‐stimulated arachidonic acid release by cells overexpressing type IIA secretory phospholipase A2
Authors: Kambe Terumi , Murakami Makoto , Kudo Ichiro
Published: July 2002

By analyzing human embryonic kidney 293 cell transfectants stably overexpressing various types of phospholipase A2 (PLA2), we have shown that polyunsaturated fatty acids (PUFAs) preferentially activate type IIA secretory PLA2 (sPLA2‐IIA)‐mediated arachidonic acid (AA) release from interleukin‐1 (IL‐1)‐stimulated cells. When 293 cells prelabeled with [3H]AA were incubated with exogenous PUFAs in the presence of IL‐1 and serum, there was a significant increase in [3H]AA release (in the order AA>linoleic acid>oleic acid), which was augmented markedly by sPLA2‐IIA and modestly by type IV cytosolic PLA2 (cPLA2), but only minimally by type VI Ca2+‐independent PLA2, overexpression. Transfection of cPLA2 into sPLA2‐IIA‐expressing cells produced a synergistic increase in IL‐1‐dependent [3H]AA release and subsequent prostaglandin production. Our results support the proposal that prior production of AA by cPLA2 in cytokine‐stimulated cells destabilizes the cellular membranes, thereby rendering them more susceptible to subsequent hydrolysis by sPLA2‐IIA.
Curcumin mediated apoptosis in AK‐5 tumor cells involves the production of reactive oxygen intermediates
Authors: Bhaumik Sraboni , Anjum Rana , Rangaraj Nandini , Pardhasaradhi B.V.V , Khar Ashok
Published: July 2002

Curcumin, the active ingredient of the rhizome of Curcuma longa has anti‐inflammatory, antioxidant and antiproliferative activities. Although its precise mode of action remains elusive, studies have shown that chemopreventive action of curcumin might be due to its ability to induce apoptosis in cancer cells. Curcumin was shown to be responsible for the inhibition of AK‐5 tumor (a rat histiocytoma) growth by inducing apoptosis in AK‐5 tumor cells via caspase activation. This study was designed to investigate the mechanism leading to the induction of apoptosis in AK‐5 tumor cells. Curcumin treatment resulted in the hyperproduction of reactive oxygen species (ROS), loss of mitochondrial membrane potential (Δψm) and cytochrome c release to the cytosol, with the concomitant exposure of phosphatidylserine (PS) residues on the cell surface. This study suggests redox signalling and caspase activation as the mechanisms responsible for the induction of curcumin mediated apoptosis in AK‐5 tumor cells.
Structural and enzymatic characterization of human recombinant GDP‐D‐mannose‐4,6‐dehydratase
Authors: Bisso Angela , Sturla Laura , Zanardi Davide , De Flora Antonio , Tonetti Michela
Published: July 2002

GDP‐D‐mannose‐4,6‐dehydratase (GMD) is the key enzyme in the ‘de novo’ pathway of GDP‐L‐fucose biosynthesis. The reported cDNA sequences for human GMD predict three forms of different length, whose ‘in vivo’ occurrence and molecular properties are completely undefined. Here, we report the expression in Escherichia coli and the properties of each native recombinant GMD form. Only the 42 kDa long GMD (L‐GMD) and the 40.2 kDa (M‐GMD) forms were recovered as soluble functional proteins, while the 38.7 kDa form, short GMD (S‐GMD), lacking an N‐terminal domain critical for dinucleotide binding, was inactive and formed a precipitate. Both L‐GMD and M‐GMD are homodimers and contain 1 mol of tightly bound NADP+. Their kinetic properties (K
m, K
cat) are apparently identical and both forms are non‐competitively feedback‐inhibited by GDP‐L‐fucose to a similar extent. M‐GMD is the predominant enzyme form expressed in several human cell lines. These data seem to suggest that modulation of the ‘de novo’ pathway of GDP‐L‐fucose biosynthesis involves mechanisms other than differential ‘in vivo’ expression of GMD forms.
Expression of endothelin‐1 in macrophages and mast cells in hyperplastic human tonsils
Authors: Li Zhiyuan , Niwa Yasuharu , Rokutan Kazuhito , Nakaya Yutaka
Published: July 2002

In situ hybridization and immunohistochemical examinations of surgically resected specimens of human hyperplastic tonsils showed that macrophages in the germinal center and mast cells in the parafollicular and interfollicular areas expressed the transcript and protein of endothelin (ET)‐1, but not ET‐2 and ET‐3. The macrophages appeared to be activated, since they possessed significant amounts of inducible nitric oxide synthase. None of these expressions was observed in normal tonsil. Our results suggest that the over‐production of ET‐1 by macrophages and mast cells may be involved in the pathogenesis of hyperplastic tonsils.
Proteolytic cleavage of β‐catenin by caspases: an in vitro analysis
Authors: Van de Craen Marc , Berx Geert , Van den Brande Ilse , Fiers Walter , Declercq Wim , Vandenabeele Peter
Published: July 2002

Cleavage of structural proteins by caspases has been associated with the severe morphological changes occurring during the apoptotic process. One of the proteins regulating the connection of the actin filament with cadherins in a cell‐cell adhesion complex is β‐catenin. During apoptosis, both an N‐terminal and a small C‐terminal part are removed from β‐catenin. Removal of the N‐terminal part may result in a disconnection of the actin filament from a cadherin cell‐cell adhesion complex. We demonstrate that caspase‐8, ‐3 and ‐6 directly proteolyse β‐catenin in vitro. However, the β‐catenin cleavage products generated by caspase‐8 were different from those generated by caspase‐3 or caspase‐6. Caspase‐1, ‐2, ‐4/11 and ‐7 did not or only very inefficiently cleave β‐catenin. These data suggest that activation of procaspase‐3, ‐6 or ‐8 by different stimuli in the cell might result in a differential proteolysis of β‐catenin.
Identification of the cDNA encoding human 6‐phosphogluconolactonase, the enzyme catalyzing the second step of the pentose phosphate pathway
Authors: Collard François , Collet Jean-François , Gerin Isabelle , Veiga-da-Cunha Maria , Van Schaftingen Emile
Published: July 2002

We report the sequence of a human cDNA encoding a protein homologous to devB (a bacterial gene often found in proximity to the gene encoding glucose‐6‐phosphate dehydrogenase in bacterial genomes) and to the C‐terminal part of human hexose‐6‐phosphate dehydrogenase. The protein was expressed in Escherichia coli, purified and shown to be 6‐phosphogluconolactonase, the enzyme catalyzing the second step of the pentose phosphate pathway. Sequence analysis indicates that bacterial devB proteins, the C‐terminal part of hexose‐6‐phosphate dehydrogenase and yeast Sol1–4 proteins are most likely also 6‐phosphogluconolactonases and that these proteins are related to glucosamine‐6‐phosphate isomerases.
The effect of iron limitation on glycerol production and expression of the isogenes for NAD+‐dependent glycerol 3‐phosphate dehydrogenase in Saccharomyces cerevisiae
Authors: Ansell Ricky , Adler Lennart
Published: July 2002

When deprived of iron, Saccharomyces cerevisiae rearranges its metabolic flux towards increased glycerol production. This work examines the role and regulation of GPD1 and GPD2, encoding two isoforms of glycerol 3‐phosphate dehydrogenase, in glycerol production during iron starvation. The two genes respond differently on transfer of cells to iron‐limited conditions. Whereas the expression of GPD2 increases about 3‐fold, that of GPD1 does not exhibit significant changes. Deletion of either GPD1 or GPD2 alters the capacity for glycerol production during iron‐limited as well as iron sufficient conditions. However, loss of function of either gene does not seem to provoke compensatory flux via the other gene product. As judged from the glycerol production, the amount produced by each single mutant adds approximately up to the level produced by the parental strain. In agreement with the pattern of expression of GPD2, this gene product was estimated to account for the bulk of the glycerol production (about 60%) during iron‐limited conditions. The strong growth inhibition caused by iron starvation was reversed by the addition of iron also for a gpd1Δgpd2Δ double deletion mutant, which is unable to produce any detectable glycerol.
Structural parsimony in endonuclease active sites: should the number of homing endonuclease families be redefined?
Authors: Kühlmann Ulrike C. , Moore Geoffrey R. , James Richard , Kleanthous Colin , Hemmings Andrew M.
Published: July 2002

Homing endonucleases are classified into four families based on active site sequence motifs. Through structural comparisons we have found structural similarities between the endonuclease domain of colicin E9, an H‐N‐H motif‐containing enzyme, and both the non‐specific nuclease from Serratia and I‐PpoI, a His‐Cys box‐containing homing endonuclease. Our comparison identifies conservation at the heart of all three enzyme active sites and so argues for a re‐classification of H‐N‐H and His‐Cys box homing endonucleases as a single family. We suggest the ‘ββα‐Me family’ of homing enzymes to reflect the three elements of secondary structure and the metal ion that define the motif.
Insulin selectively activates STAT5b, but not STAT5a, via a JAK2‐independent signalling pathway in Kym‐1 rhabdomyosarcoma cells
Authors: Storz Peter , Döppler Heike , Pfizenmaier Klaus , Müller Gertraud
Published: July 2002

The STAT multigene family of transcriptional regulators conveys signals from several cytokines and growth factors upon phosphorylation by janus kinases (JAK). Activation of STAT5 is typically mediated by JAK2, but more recent data indicate a direct activation by the insulin receptor kinase. STAT5 exists in two closely homologous isoforms, STAT5a and b. We here describe the selective tyrosine phosphorylation of STAT5b in Kym‐1 cells in response to insulin. Blocking insulin signalling by HNMPA‐(AM)3, an insulin receptor kinase inhibitor, resulted in the loss of insulin‐induced STAT5b tyrosine phosphorylation, whereas the inhibition of JAK2 by the JAK selective inhibitor tyrphostin AG490 had no effect. By contrast, in the same cells, IFNγ‐induced STAT5b activation was JAK2‐dependent, indicating that this signal pathway is functional in Kym‐1 cells. We conclude from this rhabdomyosarcoma model that STAT5b, but not STAT5a is a direct target of the insulin receptor kinase.
Channel‐mediated permeation of ammonia gas through the peribacteroid membrane of soybean nodules
Authors: Niemietz Christa M , Tyerman Stephen D
Published: July 2002

Ammonia permeability of the peribacteroid membrane (PBM) from N2‐fixing soybean nodules was measured (8×10−5 m/s) using isolated PBM in a stopped‐flow spectrofluorimeter. Ammonia (NH3) uptake into PBM vesicles was inhibited by up to 42% by HgCl2 (EC50=2.9 μM, mercaptoethanol‐reversible) and reduced by ATP pre‐incubation. The activation energy of NH3 uptake (52 kJ/mol) increased (118 kJ/mol) with HgCl2. Water transport was also HgCl2‐sensitive (EC50=52.6 μM), but increased by ATP pre‐incubation. NH3 and H2O may permeate via different pathways through Nodulin 26 or there is another protein on the PBM that is permeable to NH3.
Sclerotherapy — an adjuvant therapy to endometriosis
Authors: Chang C.‐C. , Lee H.‐F. , Tsai H.‐D. , Lo H.‐Y.
Published: July 2002

Objective: Recurrent endometriomas after medical or surgical treatmentis a difficult clinical problem for those patients who wish to perform ovulation induction. Therefore we tried to investigate the efficacy of sclerotherapy as an adjuvant management before ovulation induction to preserve more ovarian tissue for folliculogenesis in ART program. Methods: Thirty‐two patients with persistent or recurrent endometrioma after surgical or medical treatment were included in this study. Transvaginal ultrasound needle guided aspiration of the cyst followed by tetracycline instillation was performed before ovulation induction. Results: There is an encouraging clinical pregnancy rate of 34.37%. Also, there is a disappointing recurrent rate of 46.87% in 12 months follow‐up course. Conclusion: The increased interest in cost‐effective outpatient therapy and the expected difficulty in surgical treatment of recurrent endometriomas made aspiration and sclerotherapy of endometrioma an attractive option before ovulation induction.
Suppression of fibrous adhesion by proteoglycan decorin
Authors: Fukui Naoshi , Fukuda Akira , Kojima Kazunori , Nakajima Kohei , Oda Hiromi , Nakamura Kozo
Published: July 2002

Small proteoglycan decorin is known to suppress the bioactivity of TGF‐β through a competitive binding with the cell surface receptors for the cytokine. Based on this knowledge, we hypothesized that decorin could reduce the formation of fibrous adhesion, because our previous study showed the neutralizing antibody to TGF‐β1 has that effect. An intra‐articular adhesion model in the rabbit knee joint was employed in this study, and decorin was administered into the joint cavity continuously during the 4 weeks of the experiment. The results of the dose—response study demonstrated that decorin suppresses formation of fibrous adhesion in a dose‐dependent manner. When the administration of decorin was limited to shorter periods, this effect was considerably impaired and the necessity of long‐term administration was demonstrated. On the other hand, when administered together with TGF‐β1, decorin still suppressed adhesion but to a lesser extent, and it was suggested that this proteoglycan could have other significant mechanism(s) to suppress adhesion besides the neutralization of TGF‐β. Thus, the present study showed that decorin could inhibit adhesion formation by both TGF‐β dependent and independent mechanisms. Considering that decorin exists ubiquitously in the body, its administration might be a promising approach to suppress adhesion. © 2001 Orthopaedic Research Society. Published by Elsevier Science Ltd. All rights reserved.
Effect of nmda antagonists on the activity ofglutaminase and aspartate aminotransferase in thedeveloping rat cerebellum
Authors: Moran Julio , Alavez Silvestre , Rivera‐gaxiola Maritza , Valencia Antonio , Hurtado Sandra
Published: July 2002

Chronic treatment of rats from postnatal day 6 to 25 with drugs that interact with the N‐methyl‐d‐aspartate (NMDA) receptor induced a differential effect on theactivity of some enzymes involved in neurotransmitter synthesis. Two of these drugs ((5R,10S)‐ (+)‐5‐methyl‐10,11‐dihydro‐5H‐dibenzo (a,d)cyclohepten‐5,10‐iminehydrogen maleate (MK‐801) and 3‐ (2‐carboxypiperazin‐4‐yl)propyl‐1‐phosphonic acid (CPP))caused a marked reduction (20–40%) of glutaminase and aspartate aminotransferase activity inthe cerebellum. These changes were observed only at a very precise time of development (i.e. 10to 19 postnatal day). The competitive antagonist, amino phosphonovaleric acid (APV), did notaffect any of the enzymes studied at all tested ages. When animals were treated with NMDA onlya slight, but significant, increase in the activity of glutaminase was observed at 9–11 postnatal dayonly. Any of the agonists or antagonists tested significantly affected the activity of lactatedehydrogenase as compared to control animals. Histologic observations of cerebella treated withthe indicated drugs showed that only MK‐801, and CPP to a lesser extent, induced a smallreduction in the width of the internal granule layer. The body weight of animals treated withMK‐801 was clearly reduced, but only in more mature rats (>16 postnatal day), when animals didnot show any alteration in the enzymes tested. These results support the suggestion thatpresynaptic influences, particularly from glutamatergic neurons, are critical to promote cerebellargranule neurons differentiation during critical periods of the cerebellar development.
Differential control of VEGF synthesis and secretion in human glioma cells by IL‐1 and EGF
Authors: Valter Markus Maria , Wiestler Otmar Dieter , Pietsch Torsten
Published: July 2002

VEGF (vascular endothelial growth factor), one of the most potent angiogenic factors, has recently been identified as an inducer of neoangiogenesis in many tumors including gliomas. VEGF itself appears to be regulated through different pathways. Since malignant gliomas frequently show EGF receptor amplification and express IL‐1, a pivotal regulatory cytokine involved in angiogenesis, we analyzed interactions between EGF/EGF receptor and IL‐1/IL‐1 receptor and VEGF in the established glioblastoma cell lines U‐87 MG and A‐172. Basal VEGF expression was an order of magnitude higher in U‐87 MG compared to A‐172. IL‐1 caused a fast and strong increase of VEGF secretion in U‐87 MG which appeared to harbor an intracellular VEGF pool for enhanced exocytosis. The IL‐1 receptor antagonist (IL‐1‐ra) reversed this effect suggesting an IL‐1 receptor‐associated mechanism. In contrast, VEGF secretion could not be increased by exogenous IL‐1 exposure in A‐172, which apparently lacked an intracellular VEGF pool for augmented exocytosis. However, IL‐1‐ra treatment alone caused a significant reduction of basal VEGF secretion in both U‐87 MG and A‐172. This suggests that baseline secretion of VEGF involves IL‐1 receptor activation by endogenously produced IL‐1. EGF also stimulated the secretion of VEGF into the cell supernatant. However, this effect, observed in both U‐87 MG and A‐172, was delayed and only occurred following replenishment of the intracellular VEGF pool. EGF upregulated the amount of VEGF mRNA. In general, the effects of IL‐1 and EGF on VEGF were additive, suggesting independent mechanisms. Since IL‐1 appears to be involved in VEGF secretion in glial tumors through an autocrine/paracrine mechanism, recombinant human IL‐1‐ra may evolve as a new agent for anti‐angiogenic glioma therapy.
GFAP gene methylation in different neural cell types from rat brain
Authors: Barresi V. , Condorelli D.F. , Giuffrida Stella A.M.
Published: July 2002

It is generally believed that specific demethylation processes take place in the promoter of tissue‐specific genes during development. It has been suggested that hypomethylation of the −1500/−1100 domain of the 5′ flanking regulatory region of the rat glial fibrillary acidic protein gene may be specific for neuroectodermal derivatives such as neurons and astrocytes. In the present work the methylation status of one of those seven CG sites (the −1176) of the ‘neuroectoderm‐specific domain’ was analyzed. In agreement with the neuroectoderm hypothesis, the −1176 site is highly demethylated in astroglial, oligodendroglial and neuronal cells, but heavily methylated in microglial and fibroblast cells. The three different glial population are derived from the same tissue (cerebral hemispheres of newborn rats) but have a different embryological origin: oligodendrocytes and astrocytes originate from neuroectoderm, while microglia is of mesodermal origin.

It is not clear if GFAP‐negative neuronal cells maintain such demethylation in the advanced stage of maturation or if they undergo a second phase of de novo methylation. In order to clarify this point we used a subcellular fractionation method which allowed us to separate two different nuclear populations from adult rat cerebral hemispheres: one enriched in neuronal nuclei (called N1) and the other enriched in glial nuclei (N2). A higher methylation level of the −1176 site was detected in the N1 fraction, suggesting the GFAP gene undergo a de novo methylation process during neuronal maturation. This observation is in agreement with recent results showing a de novo methylation of the −1176 site during postnatal brain development. We hypothesize that a DNA demethylation process takes place in neuroectodermal precursor cells and that the −1176 site persists demethylated at the earlier stages of neuronal differentiation (immature neurons) and becomes fully methylated at more advanced stages of differentiation.
Effectiveness of CEO pay‐for‐performance
Authors: Mishra Chandra S , McConaughy Daniel L , Gobeli David H
Published: July 2002

Firm performance has a generally positive, but diminishing relationship with the level of CEO pay‐for‐performance sensitivity to stock returns, consistent with the tradeoffs between incentives and risk sharing that underlie the use of pay‐for‐performance. Two moderating risk variables capture this tradeoff and significantly shape the pay‐for‐performance relationship: a firm's business risk and the standard deviation of its stock returns. At higher levels of pay‐for‐performance sensitivity, the future performance of higher risk firms is more negatively related to sensitivity than for lower risk firms. Our results support the notion that CEO risk aversion limits the benefits from incentive pay, and that when too much risk is placed on the CEO, firm performance suffers. Compensation managers should take these results into account when making changes in CEO pay‐for‐performance plans.
Shape‐Memory Polymers for Biomedical Applications
Authors: Delaey Jasper , Dubruel Peter , Van Vlierberghe Sandra
Published: June 2020

One of the most promising fields for shape‐memory polymers is the biomedical field. Shape‐memory polymers that can be triggered by various (physiological) conditions such as temperature and moisture have been reported, as well as remotely triggerable shape‐memory polymers that make use of electromagnetic fields, ultrasound, etc. These polymers have shown great promise in in vitro studies as well as in early in vivo studies. Their biocompatibility, and in some cases biodegradability, renders them excellent candidates for minimal invasive surgery and for the design of triggerable biomedical devices. The current review provides a nonexhaustive overview of recent developments realized throughout the last decade in the field of shape‐memory polymers serving specific biomedical applications while considering relevant triggers and biocompatible chemistries.
Low genetic diversity and limited genetic structure across the range of the critically endangered Mexican howler monkey (Alouatta palliata mexicana)
Authors: Melo‐Carrillo Adrián , Dunn Jacob C. , Cortés‐Ortiz Liliana
Published: June 2020

Genetic diversity provides populations with the possibility to persist in ever‐changing environments, where selective regimes change over time. Therefore, the long‐term survival of a population may be affected by its level of genetic diversity. The Mexican howler monkey (Alouatta palliata mexicana) is a critically endangered primate restricted to southeast Mexico. Here, we evaluate the genetic diversity and population structure of this subspecies based on 83 individuals from 31 groups sampled across the distribution range of the subspecies, using 29 microsatellite loci. Our results revealed extremely low genetic diversity (HO = 0.21, HE = 0.29) compared to studies of other A. palliata populations and to other Alouatta species. Principal component analysis, a Bayesian clustering method, and analyses of molecular variance did not detect strong signatures of genetic differentiation among geographic populations of this subspecies. Although we detect small but significant FST values between populations, they can be explained by a pattern of isolation by distance. These results and the presence of unique alleles in different populations highlight the importance of implementing conservation efforts in multiple populations across the distribution range of A. p. mexicana to preserve its already low genetic diversity. This is especially important given current levels of population isolation due to the extreme habitat fragmentation across the distribution range of this primate.
Organocatalytic Enantioselective Synthesis of Chiral Allenes: Remote Asymmetric 1,8‐Addition of Indole Imine Methides
Authors: Li Xingguang , Sun Jianwei
Published: June 2020

An organocatalytic enantioconvergent synthesis of chiral tetrasubstituted allenes is disclosed. With suitable chiral phosphoric acid catalysts, a range of racemic indole‐substituted propargylic alcohols reacted with nucleophiles to provide efficient access to a series of enantioenriched allenes with high enantioselectivities. Control experiments suggested a mechanism involving remotely controlled asymmetric 1,8‐addition of the in situ generated indole imine methide via a bifunctional transition state.
Biokatalyse: Enzymatische Synthese für industrielle Anwendungen
Authors: Wu Shuke , Snajdrova Radka , Moore Jeffrey C. , Baldenius Kai , Bornscheuer Uwe T.
Published: June 2020

Die Biokatalyse hat in verschiedenen Einsatzgebieten Anwendung als Alternative zur chemischen Katalyse gefunden, wobei die herausragendsten Beispiele die Herstellung chiraler Verbindungen für Pharmazeutika und für die Riechstoff‐ und Aromen‐Industrie sind. Zunehmend werden Biokatalysatoren im großen Maßstab genutzt, um Spezial‐ und sogar Bulkchemikalien herzustellen. Dieser Aufsatz soll lehrreiche Beispiele in diesem Gebiet mit besonderem Augenmerk auf skalierbare chemische Verfahren aufzeigen. Es werden Möglichkeiten und Grenzen enzymatischer Synthesen diskutiert, und es wird ein Ausblick auf neu aufstrebende Enzymklassen gegeben.
Ein Schwefelmonoxid‐Addukt eines frustrierten Sn/P‐Lewis‐Paares
Authors: Holtkamp Philipp , Glodde Timo , Poier Dario , Neumann Beate , Stammler Hans‐Georg , Mitzel Norbert W.
Published: June 2020

Die Reaktion des geminalen frustrierten Lewis‐Paares (F5C2)3SnCH2P(tBu)2 (1) mit N‐Sulfinylanilin PhNSO ergab das erste Schwefelmonoxid‐Addukt eines Hauptgruppenmetalls (F5C2)3SnCH2P(tBu)2⋅SO (2). Es enthält einen SnCPSO‐Ring. Das zweite Produkt ist ein Phenylnitren‐Addukt von 1. Die überraschende Stabilität von 2 wurde mit denen der bisher unzugänglichen O2‐ und S2‐Addukte von 1 verglichen. Versuche, diese aus 1 und den elementaren Chalkogenen (O2, S8, Se∞, Te∞) herzustellen, führten zu viergliedrigen SnCPE‐Ringsystemen. Quantenchemische Untersuchungen von 2 zeigen den Beitrag der Bindungspolarität der SO‐Einheit zur Stabilisierung von 2.
Total Synthesis of (+)‐Cornexistin
Authors: Steinborn Christian , Wildermuth Raphael E. , Barber David M. , Magauer Thomas
Published: June 2020

Herein, we describe the first total synthesis of (+)‐cornexistin as well as its 8‐epi‐isomer starting from malic acid. The robust and scalable route features a Nozaki–Hiyama–Kishi reaction, an auxiliary‐controlled syn‐Evans‐aldol reaction, and a highly efficient intramolecular alkylation to form the nine‐membered carbocycle. The delicate maleic anhydride moiety of the nonadride skeleton was constructed from a β‐keto nitrile. The developed route enabled the synthesis of 165 mg (+)‐cornexistin.
Ir‐Catalyzed Enantioconvergent Synthesis of Diversely Protected Allenylic Amines Employing Ammonia Surrogates
Authors: Glatz Fabian , Petrone David A. , Carreira Erick M.
Published: June 2020

The first iridium catalyzed, enantioconvergent amination of allenylic carbonates is reported. This process utilizes various commercially available carbamates and sulfonamides to generate allenylic amines including commonly employed protected groups (Boc, Fmoc, Cbz, Ts, Ns) in 62–82 % yield and 87–98 % ee. The products generated through this scalable procedure serve as effective linchpins for the rapid, enantiospecific synthesis of a wide range of complex structures.
Graphdiyne:Structure of Fluorescent Quantum Dots
Authors: Guo Jie , Guo Mengyu , Wang Fuhui , Jin Weiyue , Chen Chunying , Liu Huibiao , Li Yuliang
Published: June 2020

Graphdiyne (GDY) as an emerging two‐dimensional carbon allotrope exhibits excellent performance in energy chemistry, catalytic chemistry, optoelectronics, electronics, etc. because of the unique structure combining an sp‐ and sp2‐hybrid carbon network. However, the poor solubility of pristine GDY is a major obstacle to its applications in many fields. Proposed here is a facile strategy to control the preparation of GDY quantum dots (GDY‐Py QDs), in which pyrene groups are covalently linked to GDY by using a Sonogashira cross‐coupling reaction. The as‐prepared GDY‐Py QDs, with an average diameter of about 3±0.1 nm, show superior dispersibility in many organic solvents and water. The GDY‐Py QDs display not only bright fluorescent with a high relative quantum yield (QY) of 42.82 %, but they are also well‐behaved as contrast agents in cell imaging. The GDY‐Py QDs are bestowed with high stability and non‐cytotoxicity, and exhibit long fluorescent times, and have potential for optical imaging and biomedical applications.
An Adduct of Sulfur Monoxide to a Frustrated Sn/P Lewis Pair
Authors: Holtkamp Philipp , Glodde Timo , Poier Dario , Neumann Beate , Stammler Hans‐Georg , Mitzel Norbert W.
Published: June 2020

The geminal frustrated Lewis pair (F5C2)3SnCH2P(tBu)2 (1) reacted with N‐sulfinylaniline PhNSO to afford the first sulfur monoxide adduct of a main group metal, (F5C2)3SnCH2P(tBu)2⋅SO (2), which contains a SnCPSO ring. The second product is a phenylnitrene adduct of 1. The surprising stability of 2 was compared with the stabilities of the so far inaccessible O2 and S2 adducts of 1. Attempts to prepare these from 1 and the elemental chalcogens (O2, S8, Se∞, Te∞) led to four‐membered SnCPE ring systems. Quantum‐chemical investigations of 2 demonstrate the bond polarity of the SO unit to stabilize 2.