All issues
Volume 130 (2001)
- Issue 6 Pages 721-
- Issue 5 Pages 575-
- Issue 4 Pages 471-
- Issue 3 Pages 319-
- Issue 2 Pages 169-
- Issue 1 Pages 1-
Volume 130, Issue 2
Displaying 1-20 of 20 articles from this issue
- |<
- <
- 1
- >
- >|
-
Kazuyuki Nakajima, Shinichi Kohsaka2001 Volume 130 Issue 2 Pages 169-175
Published: 2001
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSMicroglia are resident monocyte-lineaged cells in the brain. Their characteristic feature is that they react to injury and diseases of the brain and become morphologically and functionally activated. Although some triggar molecules which activate microglia are predicted to be released from injured or affected cells, such molecules have not yet been identified. The main role of activated microglia is believed to be in brain defense, as scavengers of dead cells, and as immune or immunoeffector cells. Recent biochemical and neurobiological studies have further indicated that they significantly affect the pathological state and/or regulate the regenerative state and remodeling of the brain by producing a variety of biologically active molecules including cytotoxic and neurotrophic molecules.View full abstractDownload PDF (572K) -
Kiyonao Sada, Tomoko Takano, Shigeru Yanagi, Hirohei Yamamura2001 Volume 130 Issue 2 Pages 177-186
Published: 2001
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSNon-receptor type of protein-tyrosine kinase Syk contains 2 Src homology 2 (SH2) domains in tandem and multiple autophosphorylation sites. Syk is activated upon binding of tandem SH2 domains to immunoreceptor tyrosine-based activating motif (ITAM) and plays an essential role in lymphocyte development and activation of immune cells. Syk is critical for tyrosine phosphorylation of multiple proteins which regulate important pathways leading from the receptor, such as Ca2+ mobilization and mitogen-activated protein kinase (MAPK) cascades. Recent findings reveal that expression of Syk appears to be involved in a wide variety of cellular functions and pathogenesis of malignant cancer. These observations have demonstrated that Syk is a key molecule that controls multiple physiological functions in cells.View full abstractDownload PDF (3647K) -
Izuho Hatada, Sumiyo Morita, Yayoi Obata, Yusuke Sotomaru, Masafumi Sh ...2001 Volume 130 Issue 2 Pages 187-190
Published: 2001
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSSystematic screening of differentially expressed genes among androgenetic, partheno-genetic, and normal embryos by means of fluorescent differential display revealed five imprinted genes. One of them, named Rian, was expressed exclusively from the maternal allele and was closely linked to an imprinted gene, Meg3(Gt12), mapped to the distal end of chromosome 12. The Rian transcript did not have any apparent open reading frame, and its transcript was exclusively localized to the nucleus.View full abstractDownload PDF (1530K) -
Nathaniel J. Cosper, Robert A. Scott, Hiroyuki Hori, Takeshi Nishino, ...2001 Volume 130 Issue 2 Pages 191-198
Published: 2001
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSNitric oxide synthase (NOS) catalyzes the conversion of L-arginine to citrulline and nitric oxide through two stepwise oxygenation reactions involving N--hydroxy-L-arginine, an enzyme-bound intermediate. The Nω-hydroxy-L-arginine-and arginine-bound NOS ferriheme centers show distinct, high-spin electron paramagnetic resonance signals. Iron X-ray absorption spectroscopy (XAS) has been used to examine the structure of the ferriheme site in the Nω-hydroxy-L-arginine-bound full-length neuronal NOS in the presence of (6R)-5, 6, 7, 8-tetrahydro-L-biopterin. Iron XAS shows that the high-spin ferriheme sites in the Nω-hydroxy-L-arginine-and arginine-bound forms are strikingly similar, both being coordinated by the heme and an axial thiolate ligand, with an Fe-S distance of ca. 2.29 Å. Cu2+ inhibition slightly affects the spin-state equilibrium, but causes no XAS-detectable changes in the immediate ferriheme coordination environment of neuronal NOS. The structure and ligand geometry of the high-spin ferriheme in arginine-bound neuronal NOS are essentially identical to those of the Nω-hydroxy-L-arginine-bound form and only slightly affected by the divalent cation inhibitor of consitutive NOS.View full abstractDownload PDF (712K) -
Hayato Umekawa, Keiji Sato, Masaharu Takemura, Yoshinori Watanabe, Sat ...2001 Volume 130 Issue 2 Pages 199-205
Published: 2001
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe protein B23 is a major nucleolar phosphoprotein comprising two isoforms, B23.1 and B23.2, which differ only in their carboxyl-terminal short sequences, the N-terminal 255 residues being identical in both forms. Both B23.1 and B23.2 stimulated immunoaf-finity-purified calf thymus DNA polymerase a in a dose-dependent manner. The stimula-tory effect of protein 1323.1, the longer isoform, was found to be 2-fold greater than that of B23.2. Purified DNA polymerase α bound tightly to a protein B23.1-immobilized column, while it bound weakly to a protein B23.2-immobilized column. Surface plasmon resonance studies by BlAcore further showed that protein B23.1 bound to the DNA polymerase α-(dA)•(dT) complex more tightly than did protein B23.2. The protein B23 iso-forms appear to interact directly with the DNA polymerase α protein and not through the bound nucleic acid. These observations indicated that protein B23 physically bound to the DNA polymerase α and stimulated the enzyme activity. Product analyses showed that protein B23 greatly enhanced the reaction both in amount and length of product DNA, whereas it did not significantly alter the processivity of polymerization. In contrast, protein B23 effectively protected DNA polymerase α from heat inactivation. These results suggest that protein 1423 stabilizes DNA polymerase α that is detached from product DNA, allowing the enzyme to be recruited for further elongation. Moreover, experiments using various C-terminal deletion mutants of protein B23 indicated that 12 amino acids at the C-terminal end of B23.1, which are absent in B23.2, may be essential for the full stimulation of the DNA polymerase α.View full abstractDownload PDF (2026K) -
Susumu Ohkouchi, Keiko Nishio, Mineko Maeda, Kiyotaka Hitomi, Hiroyuki ...2001 Volume 130 Issue 2 Pages 207-215
Published: 2001
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSPenta-EF-hand (PEF) proteins such as ALG-2 (apoptosis-linked gene 2 product) and the calpain small subunit are a newly classified family of Ca2+-binding proteins that possess five EF-hand-like motifs. We identified two mutually homologous PEF proteins, designated DdPEF-1 and DdPEF-2 (64% amino acid residue identities), in the cellular slime mold Dictyostelium discoideum. Both PEF proteins showed a higher similarity to mammalian ALG-2 and peflin (Group I PEF proteins) than to calpain and sorcin subfamily (Group II PEF proteins) in the first EF-hand (EF-1) regions. Northern blot analyses revealed that DdPEF-1 and DdPEF-2 were constitutively expressed throughout development of Dietyostelium, but their levels of expression were developmentally regulated. In situ hybridization analyses demonstrated that DdPEF-1 was expressed in both the ante-rior prestalk and the posterior prespore regions of the tipped aggregate, slugs and early culminants. On the other hand, DdPEF-2 was dominantly expressed in the anterior tip region of these multicellular structures. Both PEF proteins were detected as 22-23-kDa proteins in soluble fractions in the presence of EGTA but in particulate fractions in the presence of Ca2+ by Western blotting using specific monoclonal antibodies. Together with the finding of PEF-like sequences in DNA databases of plants, fungi and protists, our results strongly suggest that Group I PEF proteins are ubiquitously present in all eukaryotes and play important roles in basic cellular functions.View full abstractDownload PDF (2653K) -
Miho Matsumoto, Yoshihide Miyake, Mana Nagita, Hiroki Inoue, Daiya Shi ...2001 Volume 130 Issue 2 Pages 217-225
Published: 2001
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSWe surveyed proteins capable of binding to the cytoplasmic domain of Na+/H+ exchanger (NHE)1 in a rat brain cDNA library with the yeast two-hybrid system. One clone obtained coded for a protein reported previously as a human calcineurin homologous protein (CHP). Since CHP is homologous to the regulatory subunit B of calcineurin, we expected a possible interacting partner of CHP like the catalytic subunit of calcineurin (calcineurin A), and surveyed this putative partner again with the yeast two-hybrid system. A clone thus obtained coded for a kinase, which is basically the same as that reported for human DRAK2. Overexpression of the rat homologue of DRAK2 caused apop-tosis-like cell death of NI113T3 cells, which was dependent on the kinase activity, confirming the previous result for DRAK2. The purified CHP and rat DRAK2 proteins synthesized in Escherichia coli could bind in vitro. CHP and rat DRAK2 expressed in COS-7 cells were found to be localized in the Golgi apparatus and nucleus, respectively. Some of them was also found in the membrane peripheral region. When they were coexpressed in the same cells, most of CHP moved to the nucleus where rat DRAK2 is located, suggesting in vivo interaction of these proteins. However, minor but significant fractions of both proteins were also found in the membrane peripheral region. Rat DRAK2 is expressed highly in thymus, spleen, and testis, where the apoptosis plays an important role in physiology.View full abstractDownload PDF (2001K) -
Didier Bonnet, Julie M. Stevens, Rodolphe Alves de Sousa, Marie-Agn&eg ...2001 Volume 130 Issue 2 Pages 227-233
Published: 2001
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThere is growing evidence in the literature emphasizing the significance of the post-translational modification of cysteine thiols to sulfenic acids (SOH), which have been found in a number of proteins. Crystallographic and mass spectrometric evidence has shown the presence of this group in an inactive form of the industrially important enzyme nitrile hydratase (NHase). This oxidized cysteine is unique in that it forms part of the coordination sphere of the low-spin iron III at the active site of the enzyme. The presence of this unstable sulfenic group in the active form of NHase is the subject of some controversy. To try to detect this function in NHase, we have studied the inhibitoryeffect on nitrile hydration of reagents known to react with sulfenic acids. Two NHases were studied, namely, Rhodococcus rhodochrous R312 NHase and Comamonas testosteroni NI1 NHase, and the reagents used were meta-chlorocarbonyldicyano-phenylhydrazone (m-ClCP), 7-chloro-4-nitrobenzo-2-oxa-1, 3-diazole (NBD-Cl), and 2-nitro-5-thiocyanato-benzoic acid (NTBA). Following this approach we report three novel inhibitors of NHases. In addition, we report thiocyanate reagents that can be used to monitor NHase activity spectroscopically.View full abstractDownload PDF (2016K) -
Akira Hattori, Ken Matsumoto, Shigehiko Mizutani, Masafumi Tsujimoto2001 Volume 130 Issue 2 Pages 235-241
Published: 2001
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe genomic organization of the gene encoding the human adipocyte-derived leucine aminopeptidase (A-LAP) has been determined. The gene is composed of 20 exons and 19 introns and spans approximately 47 kilobases of chromosome 5q15. The gluzincin aminopeptidase motif, the HEXXH(X)18E zinc-binding motif essential for enzymatic activity, is encoded by exons 6 and 7. A comparison of the exon/intron boundaries, together with phylogenetic analysis, shows the close relationship between A-LAP and placental leucine aminopeptidase (P-LAP)/oxylocinase, another gluzincin aminopeptidase considered to be important for the maintenance of normal pregnancy. Primer extension analysis revealed two transcriptional initiation sites. Analysis of the sequence immediately upstream of the transcriptional initiation sites revealed that the A-LAP promoter contains no canonical TATA-or CCAAT-box, but has a PyPyA+1N(T/A)PyPy initiator consensus sequence and multiple putative regulatory elements. Finally, luciferase-reporter assays revealed a functional promoter activitiy of the 5'-flanking region of the gene, and suggested that the activity is regulated in a cell type-specific manner.View full abstractDownload PDF (1193K) -
Sawako Yamashiro, Kazuhisa Kameyama, Nobuyuki Kanzawa, Toru Tainiya, I ...2001 Volume 130 Issue 2 Pages 243-249
Published: 2001
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSMimosa pudica L. rapidly closes its leaves and bends its petioles downward when mechanically stimulated. It has been suggested that the actin cytoskeleton is involved in the bending motion since both cytochalasin B and phalloidin inhibit the motion. In order to clarify the mechanism by which the actin cytoskeleton functions in the motion, we attempted to fmd actin-modulating proteins in the M. pudica plant by DNase I-affinity column chromatography. The EGTA-eluate from the DNase I column contained proteins with apparent molecular masses of 90-and 42-kDa. The 42-kDa band consisted of two closely migrating components: the slower migrating component was actin while the faster migrating components was a distinct protein. The eluate showed an activity to sever actin filaments and to enhance the rate of polymerization of actin, both in a Ca2+-dependent manner. Microsequencing of the faster migrating 42-kDa protein revealed its similarity to proteins in the gelsolin/fragmin family. Our results provide the first biochemical evidence for the presence in a higher plant of a gelsolin/fragmin family actinmodulating protein that severs actin filament in a Ca2+-dependent manner.View full abstractDownload PDF (1737K) -
Yoshiya Ikawa, Kenji Nohmi, Shota Atsumi, Hideaki Shiraishi, Tan Inoue2001 Volume 130 Issue 2 Pages 251-255
Published: 2001
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSNatural RNAs often contain terminal loops consisting of GNRA (N=A, G, C, U; R=A, G) and their receptors, which bind to the loops via long-range RNA-RNA interactions. Among several known receptors, two characteristic structural elements have been identified that are termed the 11-nt motif (CCUAAG-UAUGG) and IC3 motif (CCCUAAC-GAGGG). These two motifs that share a similar secondary structure have been shown to exhibit distinctively different binding specificities. The 11-nt motif recognizes a GAAA loop with highest specificity among the known receptors, whereas the IC3 motif distinguishes GAAA from other GNRA loops less stringently than any other receptors. To identify the elements in the receptors that determine the binding specificity, a series of chimeric receptors derived from the two motifs were prepared and their properties were examined. We identified characteristic base-pairs and a particular U residue in the receptors as such elements by means of a gel mobility shift assay that evaluates the degree of the tetraloop-receptor interaction. The relationship between the elements and the specificity is discussed together with a model that describes a possible evolutional linkage between the two receptors.View full abstractDownload PDF (1727K) -
Yasuo Yamauchi, Yukinori Ejiri, Toshio Sugimoto, Kuni Sueyoshi, Yoshik ...2001 Volume 130 Issue 2 Pages 257-261
Published: 2001
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSWe purified a glutamyl endopeptidase that is a major foliar endopeptidase in cucumber. The endopeptidase had a molecular mass of 400 kDa, consisted of four subunits of 97 kDa, and was inactivated by SH-modifying reagents. Its optimum pH and optimum temperature were 8.0 and 30-37°C, respectively. An internal amino acid sequence of the endopeptidase was highly homologous to a partial sequence of unidentified proteins deduced from genetic information for Arabidopsis thaliana, soybean and rice, but not to the sequences of bacterial glutamyl endopeptidases or animal proteases. Therefore, the unidentified proteins might be glutamyl endopeptidases and be widely distributed only among plant species. The activity of the cucumber glutamyl endopeptidase was inhibited by at least three inhibitors existing in cucumber leaves. One of the inhibitors was a competitive inhibitor of 25 kDa, which did not significantly inhibit commercial endopeptidases derived from animals and microorganisms. This suggests that the cucumber glutamyl endopeptidase might be controlled by endogenous inhibitors in vivo.View full abstractDownload PDF (1666K) -
Yasuhiro Horibata, Hideyoshi Higashi, Makoto Ito2001 Volume 130 Issue 2 Pages 263-268
Published: 2001
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSEndoglycoceramidase (EGCase: EC 3.2.1.123) is an enzyme capable of cleaving the glycosidic linkage between oligosaccharides and ceramides in various glycosphingolipids. We report here transglycosylation and reverse hydrolysis reactions of EGCase from the jellyfish Cynaea nozakii. Various alkyl-GM1 oligosaccharides (alkyl-II3NeuAcGgOse4) were synthesized when GM1 ganglioside was treated with the EGCase in the presence of 1-alkanols. Among various 1-alkanols tested, methanol was found to be the most preferential acceptor, followed by 1-hexanol and 1-pentanol. GM1 was the best donor, followed by GD1b and GT1b, when methanol was used as an acceptor. However, neither globoside nor glucosylceramide was utilized by the enzyme as a donor substrate. The enzyme transferred oligosaccharides from various glycosphingolipids to NBD-ceramide, a fluorescent ceramide, producing NBD-labeled glycosphingolipids. In addition to the transglycosylation reaction, the enzyme catalyzed the reverse hydrolysis reaction; lactose was condensed to ceramide to generate lactosylceramide in the presence of the enzyme. These results indicate that the jellyfish enzyme will facilitate the synthesis of various neoglycoconjugates and glycosphingolipids.View full abstractDownload PDF (2634K) -
Kenji Soejima, Noriko Mimura, Hiroshi Yonemura, Hiroshi Nakatake, Taka ...2001 Volume 130 Issue 2 Pages 269-277
Published: 2001
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSHuman recombinant prethrombin-2 was produced in Eseherichia coli. The expressed prethrombin-2 formed intracellular inclusion bodies from which the protein was refolded by a simple one-step dilution process in buffer consisting of 50 mM Tris-HC1, containing 20 mM CaCl2, 500 mM NaCl, 1 mM EDTA, 600 mM arginine, 1 mM cysteine, 0.1 mM cystine10% (v/v) glycerol, and 0.2% (w/v) Brij-58 at pH 8.5. After refolding, prethrombin-2 was purified by hirudin-based COOH-terminal peptide affinity chromatography, and then activated with Echis carinatus snake venom prothrombin activator (ecarin). The activated protein, α-thrombin, was then tested for several activities including activity toward chromogenic substrate, release of fibrinopeptide A from fibrinogen, activation of protein C, and thrombin-activatable fibrinolysis inhibitor, reactivity with antithrombin, clotting activity, and platelet aggregation. The kinetic data showed no differences in activity between our recombinant α-thrombin and plasma-derived α-thrombin. The yield of refolded recombinant human prethrombin-2 was about 4-7% of the starting amount of solubilized protein. In addition, the final yield of purified refolded protein was 0.5-1%, and about 1 mg of recombinant prethrombin-2 could be isolated from 1 liter of E. coli cell culture.View full abstractDownload PDF (1333K) -
Tadanobu Takahashi, Yasuo Suzuki, Daisuke Nishinaka, Nana Kawase, Yuki ...2001 Volume 130 Issue 2 Pages 279-283
Published: 2001
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe majority of influenza A viruses isolated from wild birds, but not humans, can replicate in the duck intestinal tract. Here we demonstrate that all duck isolates tested universally retain sialidase activities under low pH conditions independent of their neuraminidase (NA) subtypes. In contrast, the sialidase activities of most isolates from humans and pigs practically disappear below pH 4.5, with the exception of four human pandemic viruses isolated in 1957 and 1968. Sequence comparisons among duck, human, and swine N2 NA subtypes indicate that amino acids at positions 153, 253, 307, 329, 344, 347, 356, 368, 390, and 431 may be associated with the low pH stability of duck and human pandemic N2 NAs. This finding suggests that the low pH stability of duck influenza A virus NA may be a critical factor for replication in the intestinal tract through the digestive tract of ducks, and that the properties of NAs are important for understanding the epidemiology of the influenza virus.View full abstractDownload PDF (360K) -
Toshiyuki Matsunaga, Takanori Nakajima, Masaru Sonoda, Iwao Koyama, Sh ...2001 Volume 130 Issue 2 Pages 285-293
Published: 2001
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSPeroxynitrite has been implicated in the oxidative modification of low-density lipoprotein (LDL) particles, and nitrotyrosine residues in the LDL have been detected in atherosclerotic plaques. Studies have suggested that lipoproteins modified by peroxynitrite lead to the onset of atherosclerotic vascular disease. We therefore prepared in vitro lipoproteins oxidatively modified by peroxynitrite (NO2-lipoprotein) and investigated the effect of NO2-lipoprotein on the viability of cultured endothelial cells. After exposure of a high-density lipoprotein (HDL) to peroxynitrite, some intermolecular complexes of apolipoproteins in HDL were detected on immunoblotting with monoclonal antibodies against apolipoprotein AI and AII suggesting that nitration of HDL by peroxynitrite causes intermolecular cross-linking of the apolipoproteins in the particles. Treatment with 1 mM peroxynitrite increased the 3-nitrotyrosine level to 28.5 mmol/mol of tyrosine residues in the prepared NO2-HDL, as quantitated by HPLC, and the amount in NO2-lipoprotein depended on the peroxynitrite concentration. HDL exhibited a shorter lag phase and the reaction plateaued more rapidly than that with LDL. To clarify whether or not NO2-lipoproteins affect the function of endothelial cells, we first examined the viability of cultured human aortic endothelial cells (HAECs) exposed to NO2-lipopro-teins. Incubation with either NO2-HDL or NO2-LDL significantly reduced the HAEC via-bility at 72h. The results of RT-PCR and Western blotting showed that NO, -HDL markedly suppressed at 48h not only the expressed levels of mRNA and protein but also the activity of catalase in HAECs. In contrast, NO2-LDL significantly reduced the expres-sion and activity of Cu2+, Zn2+-superoxide dismutase (CuZn-SOD) in the cells. Neither NO2-HDL nor NO2-LDL interfered with nitric oxide production or expression of cycloox-ygenases and NADPH oxidase in HAECs. Increased radical production in NO2-lipopro-tein-treated HAECs implied that reactive oxygen species such as superoxide anions and hydroxyl radicals may contribute to the mechanism of the toxic effect induced in endo-thelial cells by NO2-lipoprotein. Overall, NO2-lipoprotein may lead to deterioration of the vascular function through these endothelial cell responses.antioxidant enzyme, endothelial cell, lipoprotein, peroxynitrite, reactiveoxygen species.View full abstractDownload PDF (2668K) -
Eiji Kurimoto, Takushi Harada, Ayako Akiyama, Tomoya Sakai, Koichi Kat ...2001 Volume 130 Issue 2 Pages 295-297
Published: 2001
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSSince in vitro refolding of pepsin has long been attempted without success, it has been suspected that pepsin has no intrinsic refolding ability. In the present study, in order to eliminate unfavorable intermolecular interactions bringing about aggregation and autoproteolysis, we immobilized pepsin onto agarose beads. This technique enabled us to search extensively for appropriate refolding conditions without limitation of the refolding period. Renaturation of immobilized pepsin was observed exclusively at pH 3-5. This process was extremely slow and reached equilibrium after 300 h. Sixty percent of the proteolytic activity was recovered at pH 5. Addition of salts raised the recovery to 80% but had no significant effect on the refolding rate, suggesting that the salts mainly stabilize the native state of pepsin. This is the first report on the successful in vitro refolding of pepsin.immobilization, multidomain protein, pepsin, pro-peptide, protein refolding.View full abstractDownload PDF (256K) -
Masataka Takekoshi, Fumiko Maeda, Yasuko Nagatsuka, Satoshi Aotsuka, Y ...2001 Volume 130 Issue 2 Pages 299-303
Published: 2001
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSPeripheral blood was obtained from a healthy human volunteer and transformed with Epstein-Barr virus (EBV). This produced an oligoclonal cell library in culture medium that was screened by ELISA for anti-human tumor necrosis factor-α (TNFα) activity. RNA from two positive clones was applied to RT-PCR using antibody-specific primers, and the light (κ and λ) and heavy chain genes (γ and μ) were cloned into the plasmid vector pFabl-His2. The antibodies produced in Escherichia coli as Fab fragments were assayed for anti-TNFα activity utilizing ELISA. Two IgGl/κ anti-TNFα antibodies and two IgM/κ anti-TNFα antibodies were isolated. DNA sequence analysis showed that the VL and VII gene families of IgM and IgG were the same. Both the antibodies showed almost the same activity on ELISA-testing. Ten clones randomly selected from light (κ and λ) and heavy (γ and μ) chain genes in the oligoclonal cell library 1D5 were sequenced, and each gene (κ, λ, γ, and μ) was found to be composed of one to three different genes. These data support the conclusion that the cell clone is oligoclonal at the molecular level.View full abstractDownload PDF (453K) -
Masahiro Kawahara, Hiroshi Ueda, Kouhei Tsumoto, Izuini Kumagai, Walt ...2001 Volume 130 Issue 2 Pages 305-312
Published: 2001
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSWe report a strategy for generating efficient signal transduction with unnatural heterologous receptor combinations. As previously described [Ueda, H., Kawahara, M. et al. (2000) J. Immunol. Methods 241, 159-170], chimeric receptors composed of the VH/VL domains of anti-hen egg lysozyme antibody HyHEL-10 and N-terminally truncated erythropoietin receptor (EpoR) can be activated by lysozyme. When the cytoplasmic domains of these receptors were substituted with one derived from gp130, IL-3 dependent Ba/F3 cells expressing both VH-gp130 and VL-gp130 grew dose-dependently when given lysozyme without IL-3. However, cells expressing the heterologous pair of VH-gp130 and VL-EpoR also showed more efficient and stricter lysozyme-dependent proliferation in the absence of IL-3, indicating this combination is as an efficient and strict signal transducer as wild-type EpoR. The immunoprecipitation data indicated the existence of a preformed VH-gp130 and VL-EpoR heterodimer in the absence of lysozyme, suggesting the crucial role of a receptor conformational change in signal triggering as well as wildtype EpoR and gp130. Phosphorylation of JAK2, STAT3, and STATS was observed upon the addition of lysozyme, suggesting the activation of both EpoR- and gp130-derived signals.View full abstractDownload PDF (2812K) -
Yumiko Tsuji, Tomohisa Aoyama, Koh Takeuchi, Ko-ichi Homma, Hideo Taka ...2001 Volume 130 Issue 2 Pages 313-318
Published: 2001
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSPreviously, we purified a serine protease with a molecular mass of 26 kDa that exhibits potent antibacterial activity from a pupal extract of Sarcophaga peregrina (flesh fly). We divided this protease into 12 peptides and examined their antibacterial activity. A peptide corresponding to residues 155 to 174 (peptide 9) was found to exhibit antibacterial activity comparable to that of the 26-kDa protease. When Escherichia coli was treated with peptide 9, the permeability of both the outer and inner membranes increased, and substrates for β-lactamase and β-galactosidase entered the cells, but β-galactosidase did not leak out of the cells under these conditions. It was suggested that residues 6 to 18 of peptide 9 form an amphiphilic α-helix under hydrophobic conditions with an N-terminal basic loop and then interact with acidic phospholipids in the bacterial membranes.View full abstractDownload PDF (464K)
- |<
- <
- 1
- >
- >|