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K Shimazaki

Publications and source records attributed to K Shimazaki.

At least 109 records · Page 6Linked to original sources
PubMed1993

Analysis of human and bovine milk lactoferrins by Rotofor and chromatofocusing.

1. Isoelectric points of human and bovine lactoferrins were evaluated by Rotofor and chromatofocusing analysis. 2. By Rotofor, the isoelectric value of human lactoferrin fraction was determined at 8.7 and that of bovine lactoferrin at 8.8. 3. By chromatofocusing analysis, human and bovine lactoferrins showed different elution patterns. Human lactoferrin was eluted at pH 6.8-8 and bovine lactoferrin eluted at pH 8.2-8.9.

Animals↗
PubMed1993

Immunohistochemical localization of lactoferrin in bovine exocrine glands.

The localization of lactoferrin (LF) was studied in bovine exocrine glands by the peroxidase-antiperoxidase method. In general, LF immunoreactivity was demonstrated in most of the serous demilunes of the mixed glands and in some but not all acinar cells of the serous glands. LF was not detected in the acinar cells of the mucous glands. LF immunoreactive cells were seen in the nasal and tracheal glands. The lacrimal and mammary glands were stained intensely, while the sweat glands were not stained. In the major and minor salivary glands, LF immunoreactivity was demonstrated in the serous demilunes and acini of the mixed glands and in some serous cells of the pure serous parotid gland. LF immunoreactivity was not seen in the gastrointestinal tract and pancreas, but the serous gland in the lamina propria of the bile duct was stained positively. In the male reproductive tracts, the prostate and bulbourethral glands were stained positively, but the vesicular gland was negative. In the female reproductive system, the uterine and Bartholin glands, the epithelial cells of the cervix and ampulla of the uterine tube reacted positively. However, the lesser vestibular gland did not react for LF. The present findings, that LF was demonstrated in the exocrine glands located mainly in the gateways of the digestive, respiratory and reproductive systems, may morphologically support the antibacterial role of LF.

Animals↗
PubMed1993

Separation and characterization of the C-terminal half molecule of bovine lactoferrin.

The C-terminal half molecule (C lobe) of bovine lactoferrin was isolated by mild tryptic hydrolysis of lactoferrin followed by gel filtration and ion-exchange chromatography. The identity of the fragment was established by determining its N-terminal and C-terminal amino acid sequences and comparing them with the amino acid sequence of intact lactoferrin. The isoelectric point of the C lobe ranged between pH 6.2 and 6.5 as measured by isoelectric focusing on polyacrylamide gels. The circular dichroic spectrum in the range of 250 to 350 nm of the C lobe differed slightly from that of intact lactoferrin. The pattern of lectin reactivity was similar for both the C lobe and intact lactoferrin. The C lobe showed partial antigenic identity with intact lactoferrin as demonstrated by the double immunodiffusion method, and pH dependence of iron binding of C lobe is the same as that of intact lactoferrin molecule.

Amino Acid Sequence↗
PubMed1992

Ischemia-induced changes in PIP2 levels of gerbil hippocampus.

We carried out an immunohistochemical study to detect changes in phosphatidylinositol 4,5-bisphosphate (PIP2) in gerbil hippocampus at various times after transient ischemia, using an anti-PIP2 antibody. About 24 h after transient ischemia for 5 min, an increase in the immunoreactivity was observed which was restricted to the area of CA1 pyramidal neurons. On the other hand, after less severe ischemia lasting 2 min, which did not lead to neuronal death, a decrease in PIP2 immunoreactivity was observed at about 48 h. The results indicate that levels of PIP2 following ischemia reflect dynamic changes in phosphatidylinositol (PI) turnover which may be related to neuronal degeneration.

Animals↗
PubMed1992

Purification of AMPA type glutamate receptor by a spider toxin.

A glutamate receptor was purified from Triton X-100-solubilized bovine cerebellum membranes. The purification was carried out in two steps: affinity chromatography using a spider toxin (Joro spider toxin; JSTX) immobilized on a lysine-agarose column, and a Mono Q anion exchange column. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) of the purified active fraction showed a single band with Coomassie Blue staining, which migrated with a M(r) = 130,000. The specific [3H]amino-3-hydroxy-5-methyl-isoxazole propionate ([3H]AMPA) binding activity of the affinity-purified fraction was 2095-fold higher than that of the crude soluble fraction. Lineweaver-Burk plot analysis showed a Kd of 12.7 nM [3H]AMPA in the purified fraction. The purified fraction was examined with patch-clamp recording methods in reconstituted liposomes. A glutamate-activated channel was observed and was inhibited with JSTX. The rank order of potency of agonists inducing channel currents was AMPA = glutamate greater than quisqualate much greater than kainate greater than NMDA. Thus, there is strong evidence that the 130 kDa protein is a purified component of the native AMPA type glutamate channel of bovine cerebellum.

Animals↗
PubMed1992

Different profiles of induced cotton effects of human and bovine lactoferrin by cibacron blue F3GA binding.

1. The properties of complex formation of lactoferrin with Cibacron Blue F3GA dye have been studied by circular dichroic spectral analyses. The Cotton effects were induced by the interaction of lactoferrin with the dye and occurred in the wavelength range from 300 to 450 nm. 2. The patterns of changes in circular dichroic spectra of lactoferrin induced by the dye were different in bovine and human lactoferrin. 3. Iron ions bound to lactoferrin affected the profiles of induced Cotton effects in human lactoferrin but not in bovine lactoferrin. 4. While the dye bound co-operatively to human apo-lactoferrin, such co-operativeness was not observed in iron-saturated human lactoferrin.

Animals↗
PubMed1992

Involvement of Calmodulin and Calmodulin-Dependent Myosin Light Chain Kinase in Blue Light-Dependent H Pumping by Guard Cell Protoplasts from Vicia faba L.

Signal transduction processes involved in blue light-dependent proton pumping were investigated using guard cell protoplasts from Vicia faba.N-[2-(Methylamino)ethyl]-5-isoquinolinesulfonamide, an inhibitor of cyclic AMP- and cyclic GMP-dependent protein kinases, had no effect. 1-(5-Isoquinolinesulfonyl)-2-methylpiperazine (H-7) and calphostin C, inhibitors of protein kinase C, produced slight inhibition of the blue light-dependent proton pumping. 1-[N, O-Bis(5-isoquinolinesulfonyl)-N-methyl-l-tyrosyl] -4-phenylpiperazine, a specific inhibitor of Ca(2+)/calmodulin (CaM)-dependent protein kinase II, did not inhibit the proton pumping, but 1-(5-iodonaphthalene-1-sulfonyl)-1H-hexahydro-1,4-diazepine and 1-(5-chloro-naphthalene-1-sulfonyl)-1H-hexahydro-1,4-diazepine (ML-9), inhibitors of Ca(2+)/CaM-dependent myosin light chain kinase, strongly suppressed the proton pumping. A CaM antagonist, N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide (W-7), inhibited blue light-dependent proton pumping, whereas its less active structural analog, N-(6-aminohexyl)-1-naphthalenesulfonamide (W-5), had little effect on the response. Other CaM antagonists, trifluoperazine, compound 48/80, prenylamine, and 3-(2-benzothiazolyl)-4,5-dimethoxy-N-[3-(4-phenyl-piperidinyl)- propylbenzenesulfonamide inhibited the proton pumping. In accord with these results, light-induced stomatal opening in the epidermis of Commelina benghalensis ssp. was inhibited by ML-9 and W-7, but not by H-7 and W-5. Thus, it is concluded that CaM and Ca(2+)/CaM-dependent myosin light chain kinase are the components of the signal transduction process in blue light-dependent proton pumping in guard cells.

Journal Article↗
PubMed1991

A structural study of calcium-binding equine lysozyme by two-dimensional 1H-NMR.

Since 1H-NMR spectra of the calcium bound form (holo) and the calcium free form (apo) of equine lysozyme have an overall similarity, the folded structure of apo equine lysozyme seems to be similar to the holo structure at 25 degrees C and pH 7.0, even at low ionic strengths except for subtle conformational change. However, calcium titration experiments showed that a number of resonances change by a slow exchange process. The changes saturated at one calcium ion per one lysozyme molecule, and no more change was observed by further addition of calcium ions. This shows that just one calcium ion binds to equine lysozyme. To make assignments for these changed proton resonances, two-dimensional 1H-NMR studies, correlated spectroscopy (COSY), two-dimensional homonuclear Hartmann-Hahn spectroscopy (HOHAHA) and nuclear Overhauser effect spectroscopy (NOESY) were carried out. A structural model of equine lysozyme based on the crystal structure of human lysozyme was estimated and used to assign some resonances in the aromatic and beta-sheet regions. It was possible to use some proton signals as a probe to determine the specific conformational change induced by calcium ions. The calcium binding constant KCa was estimated from calcium titration experiments in which changes in the proton signal were monitored. The log KCa value was found to be on the order of 6-7, which is in agreement with the calcium binding constant determined by fluorescence probes. This means that the protons are affected by specific calcium binding.

Amino Acids↗
PubMed1991

Structural determination of three neutral oligosaccharides in bovine (Holstein-Friesian) colostrum, including the novel trisaccharide; GalNAc alpha 1-3Gal beta 1-4Glc.

Two trisaccharides, and a pentasaccharide were obtained from bovine colostrum. Their chemical structures were determined by using methylation and 13C-NMR analyses as follows: GalNac alpha 1-3Gal beta 1-4Glc, Gal alpha-1-3Gal beta 1-4Glc, GaL beta 1-3[Gal beta 1-4GlcNAc beta 1-6]Gal beta 1-4Glc. GalNAc alpha 1-3Gal beta 1-4Glc, which was identified in this study, is a novel oligosaccharide from natural sources. Gal alpha 1-3Gal beta 1-4Glc and Gal beta 1-3[Gal beta 1-4GlcNAc beta 1-6]Gal beta 1-4Glc (lacto-N-novopentaose) have been already found in ovine colostrum, and in horse colostrum and marsupial milk, respectively.

Acetylgalactosamine↗
PubMed1991

Comparison of bovine, sheep and goat milk lactoferrins in their electrophoretic behavior, conformation, immunochemical properties and lectin reactivity.

1. The biochemical properties of bovine, goat and sheep lactoferrin were compared. Molecular weights of the three lactoferrins were estimated to be 78,000 to 80,000 as determined by SDS-PAGE. By IEF, microheterogeneity was observed for all of them. 2. Partial antigenic identity was observed between bovine lactoferrin and goat or sheep lactoferrin by immunodiffusion method. 3. CD spectra at the u.v. region of the three lactoferrins suggested their similar secondary and tertiary structural profiles. 4. Reactivities with peroxidase-conjugated lectins showed that the carbohydrate compositions of the three ruminants' lactoferrin were the same but not identical with that of human lactoferrin.

Animals↗
PubMed1991

Spider toxin and the glutamate receptors.

A neurotoxin (JSTX) was isolated from the venom of spider (Nephila clavata). JSTX blocked both the excitatory postsynaptic (EPSPs) and glutamate-induced potentials in lobster neuromuscular synapse and squid giant synapse. In mammalian central nervous system, JSTX blocked the EPSPs in CA1 pyramidal neurons resulting from stimulation of Schaffer collateral/commissure input. Pharmacological investigation showed that JSTX preferentially suppressed quisqualate/kainate receptor subtypes but was much less effective on NMDA receptor. Using synthesized spider toxins we studied the structure-activity relationship and found that the 2,4 dihydroxyphenylacetyl asparagine in the toxin structure was responsible for suppressive action, while the remaining part containing a polyamine was related to the agonist binding site with the polycationic part enhancing the toxic activity. Labeling of synthesized JSTX was used for histochemical as well as biochemical studies. Using autoradiography, 125I-JSTX-3 was found to bind at the lobster neuromuscular synapse. Histochemical study utilizing the interaction of biotinylated JSTX-3 with avidin showed specific binding of the toxin in rat cerebellum and hippocampus. JSTX-3-binding protein was purified from rat brain by affinity chromatography. SDS-PAGE of the affinity purified protein showed at least 4 bands ranging from 40 to 70 kDa.

Animals↗
PubMed1990

A histochemical study of glutamate receptor in rat brain using biotinyl spider toxin.

Joro spider toxin (JSTX), a specific blocker of glutamate receptors, was conjugated with biotin. Using the avidin-biotin complex method, specific binding sites of biotinylated JSTX were demonstrated in the cerebellum and hippocampus of the rat. In the cerebellum, strong binding of biotinyl JSTX was observed on perikarya and dendrites of the Purkinje cells with much less binding in the granular cell layer. In the hippocampus, a dense staining was observed in the pyramidal cell layer, with more heavy binding in CA3 than in other sectors of Ammon's horn. The area of distribution of biotinyl JSTX binding sites corresponded well with that of receptors preferentially activated by quisqualate.

Animals↗
PubMed1989

Effects of a spider toxin (JSTX) on hippocampal CA1 neurons in vitro.

The effect of a toxin (JSTX) obtained from Nephila clavata (Joro spider) on the CA1 pyramidal neurons of the hippocampus was studied using slice preparations. JSTX blocked the excitatory postsynaptic potentials (EPSPs) in the pyramidal neuron evoked by Schaffer collateral stimulation but was without effect on the antidromic action potentials or on the resting conductance. Depolarization induced by ionophoretic application of glutamate was readily suppressed by JSTX but aspartate-induced depolarization was much less sensitive to the toxin. Among preferential agonists activating 3 receptor subtypes for excitatory amino acids, quisqualate responses were most effectively suppressed by JSTX. Kainate responses were similarly suppressed but in some cells higher concentration of the toxin was needed to block the responses. N-methyl-D-aspartate (NMDA) responses were the least sensitive to JSTX but they were suppressed by +/- 2-amino-5-phosphonovaleric acid (APV). Long term potentiation (LTP) once it had taken place was not completely inhibited by APV. In the presence of JSTX, however, LTP was blocked and tetanic stimuli produced only a short-lived potentiation. In Mg2+ free solution, an orthodromic stimulation evoked repetitive spike responses which were superimposed on the depolarization following the initial spike. APV suppressed the depolarization and associated spikes leaving an orthodromic response which was sensitive to JSTX. The results suggest that JSTX blocks EPSPs in CA1 pyramidal neurons which are mediated by non-NMDA type receptors.

Action Potentials↗