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T Miyata

Publications and source records attributed to T Miyata.

At least 433 records · Page 24Linked to original sources
PubMed1993

Inhibition of coaggregation between Porphyromonas gingivalis and Streptococcus oralis by fibrinogen fragments.

The localization of regions of fibrinogen that inhibit coaggregation between Porphyromonas gingivalis and Streptococcus oralis was investigated. The coaggregation was inhibited by A alpha and gamma chains, but not by B beta chain. The inhibitory activity of fragment D was more potent than that of fragment E. Some cyanogen bromide-treated fragments isolated from A alpha and gamma chains including the NH2-terminal 148-207 amino acid residues of A alpha chain (A alpha 148-207) and gamma 1-78 showed inhibitory activities. A alpha 148-207 was further digested with lysyl endopeptidase. A alpha 158-176 and A alpha 192-206 which contained four and two arginine residues, respectively, retained the inhibitory activities. When the arginine residues of these two peptides were modified by phenylglyoxal, the inhibitory activities were much reduced. These findings suggest that the arginine residues of some specific regions of fibrinogen may play an important role in the inhibition of the coaggregation.

Amino Acid Sequence↗
PubMed1993

Factor IX Fukuoka. Substitution of ASN92 by His in the second epidermal growth factor-like domain results in defective interaction with factors VIIa/X.

Hemophilia B Fukuoka, a moderately severe bleeding disorder, is a naturally occurring mutant of factor IX. Plasma from our patient had 3% clotting activity even though 64% of factor IX antigen was present. The purified mutant protein was cleaved normally by factor Xla, factor VIIa-tissue factor complex, or RVV-X (factor X-activating enzyme from Russell's viper venom), yielding a two-chain factor IXa. Amino acid composition and sequence analyses of one of the lysyl endopeptidase peptides derived from factor IX Fukuoka revealed that Asn92 in the second epidermal growth factor (EGF)-like domain had been replaced by His. The active site of the factor IXa Fukuoka was normally competent for the incorporation of p-aminobenzamidine and for the hydrolysis of a synthetic substrate, N alpha-benzyloxycarbonyl-L-arginine p-nitrobenzyl ester. Factor Xa formation by factor IXa Fukuoka was only 8% of the normal factor IXa, even in the presence of polylysine, and only 0.2% of the normal in the system containing phospholipids, Ca2+, and factor VIIIa, thereby indicating a functional defect in interaction of the mutant with factors VIIIa/X. Furthermore, catalytic efficiency (kcat/Km) of factor IXa Fukuoka toward factor X in the presence of Ca2+, phospholipids, and factor VIIIa was only 2.3% of the normal factor IXa. These results suggest that an Asn-to-His substitution at position 92 in the second EGF-like domain of factor IX Fukuoka would have an untoward effect on the specific conformational state of factor IX for binding with factors VIIIa/X.

Adult↗
PubMed1993

Calorimetric investigations of phase transitions of sonicated vesicles of dimyristoylphosphatidylcholine.

The process of conversion of large multilamellar vesicles (MLVs) of dimyristoylphosphatidylcholine (DMPC) into the final state of small unilamellar vesicles (SUVs) with an increase in time length of ultrasonic irradiation was investigated by calorimetry and negative-stain electron microscopy. The process was found out to be composed of two stages depending on the primary (near 24 degrees C) and secondary (near 19 degrees C) peaks due to the gel-to-liquid crystal phase (Tm) transition, respectively; a new transition peak for the secondary Tm appears after a maximum broadening of the primary Tm peak is attained. Sonicated vesicles characterized by the primary peak of the broadest shape were observed to be about 200 nm in mean diameter and mostly four or so lamellae, and have an internal aqueous space, in contrast to sonicated SUVs (approx. 40 nm in diameter) characterized by the limiting secondary Tm peak. Thermal data associated with the Tm transition for these two sonicated vesicles were compared with that of the MLV. The enthalpy and entropy changes and cooperative units increased in the order sonicated SUV < sonicated large vesicle < MLV. Furthermore, the enthalpy changes were revealed to fairly differ between the sonicated large vesicle and SUV. Based on the effect of the annealing treatment at -5 degrees C on these vesicles the present result suggested a large contribution of the aggregation state of DMPC molecules to the enthalpy possessed by the vesicles of a gel phase temperature, which is related to the mode of the Tm transitions, primary and secondary.

Calorimetry, Differential Scanning↗
PubMed1993

Crystalline cylindrical structures of Na(+)-bound dimyristoylphosphatidylglycerol as revealed by microcalorimetry and electron microscopy.

Multilamellar vesicles of dimyristoylphosphatidylglycerol (DMPG) in an aqueous medium of 1 M NaCl at pH 7.0 converted to defined cylindrical structures of crystalline phase, on annealing of 5 degrees C of a gel phase temperature. The new crystalline phase transformed into the liquid crystal phase at a transition point (44.2 degrees C) nearly the same as that of the protonated DMPG, but with a transition enthalpy (19.2 kcal/mol) larger than that of the Ca(2+)-bound DMPG of much higher transition temperature. Electron microscopic observations revealed that a set of outer membranes (8-10 lamellae) separated from the MLV grows into helical structures and, finally, into the crystalline cylindrical structures with an increase in annealing time.

Calorimetry↗
PubMed1993

Deficiency of the GPI anchor caused by a somatic mutation of the PIG-A gene in paroxysmal nocturnal hemoglobinuria.

Paroxysmal nocturnal hemoglobinuria is an acquired hematopoietic disease characterized by abnormal blood cell populations in which the biosynthesis of the glycosylphosphatidylinositol (GPI) anchor is deficient. Deficiency of surface expressions of GPI-anchored complement inhibitors leads to complement-mediated hemolysis. Here we report that PIG-A, which participates in the early step of GPI anchor biosynthesis, is the gene responsible for paroxysmal nocturnal hemoglobinuria. Affected granulocytes and B lymphocytes had the same somatic mutation of PIG-A, indicating their clonal origin from a multipotential hematopoietic stem cell. We localized PIG-A to the X chromosome, which accounts for expression of the recessive phenotype of the somatic mutation and the fact that the same one of the multiple biosynthetic steps is affected in all patients so far characterized.

B-Lymphocytes↗
PubMed1993

The cloning of PIG-A, a component in the early step of GPI-anchor biosynthesis.

The glycosylphosphatidylinositol (GPI) anchor is a membrane attachment structure of many proteins and occurs in a wide variety of eukaryotes from yeasts to mammals. The structure of the core of the GPI anchor is conserved in protozoa and mammals and so is its biosynthetic pathway. A complementary DNA encoding a human protein termed PIG-A (phosphatidylinositol glycan-class A) was cloned. PIG-A was necessary for synthesis of N-acetylglucosaminyl-phosphatidylinositol, the very early intermediate in GPI-anchor biosynthesis.

Amino Acid Sequence↗
PubMed1993

Deficient biosynthesis of N-acetylglucosaminyl-phosphatidylinositol, the first intermediate of glycosyl phosphatidylinositol anchor biosynthesis, in cell lines established from patients with paroxysmal nocturnal hemoglobinuria.

Paroxysmal nocturnal hemoglobinuria (PNH) is a hemolytic disorder caused by a deficiency of biosynthesis of the glycosyl phosphatidylinositol (GPI) anchor, but the biochemical defect is not completely understood. In the present study, we have analyzed affected cell lines established recently from two Japanese patients with PNH. Two lines of evidence indicate that these cells do not synthesize N-acetylglucosaminyl-phosphatidylinositol, the first intermediate in the GPI anchor biosynthesis. First, somatic cell hybridization analysis using Thy-1-deficient murine thymoma cell lines with known biochemical defects as fusion partners showed that the PNH cell lines belong to complementation class A, which is known not to synthesize N-acetylglucosaminyl-phosphatidylinositol. Second, analysis of in vitro glycolipid biosynthesis demonstrated that cell lysates of these PNH cell lines in fact did not support biosynthesis of N-acetylglucosaminyl-phosphatidylinositol. Thus, we have characterized for the first time the exact biochemical defect leading to PNH.

Cells, Cultured↗
PubMed1993

Regulation of human tissue factor expression by mRNA turnover.

Tissue factor serves as the cellular receptor for circulating blood coagulation factor VII and is the principal physiological initiator of blood coagulation. Tissue factor is not normally expressed in cells that contact blood, such as endothelial cells and monocytes, but can be induced in these cells by tumor necrosis factor or tumor-promoting phorbol esters. Following induction, the human tissue factor mRNA is degraded with a half-life of approximately 0.75-1.5 h. The cellular mechanisms responsible for this rapid mRNA turnover were investigated with chimeric tissue factor.beta-globin constructs expressed in stably transfected mouse NIH/3T3 cells. These constructs were expressed with the transiently inducible c-fos promoter which eliminated the need to use transcriptional inhibitors to determine mRNA half-lives. Sequences capable of conferring rapid turnover to the normally stable beta-globin transcript were localized to the last 600 nucleotides of the tissue factor mRNA. The 3' end of this fragment is similar to previously described AU-rich mRNA destabilizing elements. Activity of the tissue factor element was dependent on its specific sequence and not simply a high AU nucleotide content. The degradation of unstable chimeric tissue factor.beta-globin mRNAs was prevented by inhibition of transcription with actinomycin D. Chimeric tissue factor.beta-globin mRNAs were superinduced by the protein synthesis inhibitor cycloheximide, and this superinduction may be due in part to stabilization of the mRNA.

3T3 Cells↗
PubMed1993

Limulus hemocyte transglutaminase. Its purification and characterization, and identification of the intracellular substrates.

To investigate further the molecular events of the intracellular coagulation cascade in limulus hemocytes, a transglutaminase (TGase), which may be involved in the formation of a stabilized gel, was purified and characterized. Through the purification procedures consisting of six steps, 1.6 mg of TGase with a specific activity of 940 amine incorporation unit/mg was obtained from 32.4 g of Tachypleus tridentatus hemocytes. The purified TGase gave a single band on SDS-polyacrylamide gel electrophoresis with a molecular mass of 86 kDa, and demonstrated mammalian-type II TGase-like enzymatic properties. The TGase activity was Ca(2+)-dependent and was inhibited by primary amines, EDTA, and SH-reagents. Moreover, two major potential substrates for TGase were identified in the hemocyte lysate by using dansylcadaverine (DCA) incorporation in the presence of 10 mM CaCl2 and 10 mM dithiothreitol. Of these protein substrates, an 80-kDa protein contained a large number of proline residues, amounting to about 22% of the total amino acids. On the other hand, an 8.6-kDa protein abundantly present in the hemocytes was characterized as a Cys-rich protein consisting of 81 amino acid residues and a calculated molecular mass of 8,671. The entire amino acid sequence of this protein was established. Also, the 8.6-kDa protein was readily cross-linked intermolecularly by TGase, forming multimers as large as pentamers. We speculate that like plasma factor XIIIa, limulus TGase and its two protein substrates in the hemocytes may play an important role in the defense of this animal against invading microorganisms.

Amino Acid Sequence↗
PubMed1993

Limulus hemocyte transglutaminase. cDNA cloning, amino acid sequence, and tissue localization.

We have evidence that the limulus (Tachypleus tri-dentatus) hemocyte transglutaminase (TGase) has a molecular mass of 86 kDa and properties of the mammalian type II TGase-like enzyme (Tokunaga, F., Yamada, M., Miyata, T., Ding, Y.-L., Hiranaga-Kawabata, M., Muta, T., Iwanaga, S., Ichinose, A., and Davie, E.W. (1993) J. Biol. Chem. 268, 252-261). We present here the cDNA and amino acid sequences, and localization of the TGase in various tissues of limulus. The cloned cDNA for TGase consists of 2,884 base pairs. An open reading frame of 2,292 base pairs encodes a sequence comprising 764 residues of the mature protein with molecular masses of 87,021 and 87,110 Da, due to two different clones. The discrepancies of nucleotides in these two clones result in 3 amino acid exchanges at positions Gly452(GGT)-Arg(CGT), Ser477(AGT)-Cys(TGT), and Ile486(ATC)-Ser(AGC), respectively. Northern blot analysis on a total RNA extracted from various tissues of limulus revealed that TGase is expressed with 3.0 kilobases of a single type of mRNA, mainly in hemocytes, hepatopancreas, and gastric tissues. Limulus TGase shows significant sequence similarity with the mammalian TGase family, as follows: guinea pig liver TGase (32.7%), human factor XIIIa subunit (34.7%), human keratinocyte TGase (37.6%), and human erythrocyte band 4.2 (23.0%). Limulus TGase has a unique NH2-terminal cationic extension of 60 residues with no homology to the NH2 termini of mammalian TGases. Based on the alignment of the amino acid sequence of limulus TGase with those of the known TGase family, a phylogenetic tree representing an evolutionary relationship among the family members was inferred by the neighbor joining method.

Amino Acid Sequence↗
PubMed1993

Characterization of neutrophil activation by repeated injection of endotoxin in rabbits. Role of neutrophils in the generalized Shwartzman reaction.

The relationship between activated neutrophils and end-organ injury in endotoxemia was studied. The function of peripheral blood neutrophils (PMNs) in rabbits with the generalized Shwartzman reaction (GSR) was compared to that of PMNs rabbits receiving a single injection of endotoxin. The following results were obtained: (1) PMNs from rabbits with the GSR demonstrated enhanced adherence to endothelial cells and increased mitochondrial ATP production; (2) the GSR did not enhance chemotaxis and oxygen radical production of PMNs; (3) a single injection of endotoxin did not cause necrosis of visceral organs; (4) in vitro detachment of endothelial cells by PMNs was increased in rabbits with the GSR; (5) in vivo administration of monoclonal antibody (mAb) against CD11b/CD18 (Mac-1) suppressed the increase in PMN adherence; and (6) hemorrhagic necrosis did not occur when mAb to Mac-1 was injected. Thus, enhanced adherence of PMNs to endothelial cells appears to play a key role in endotoxin-induced end-organ injuries in this animal model.

Animals↗
PubMed1993

Decreased activity of complement-mediated immune complex clearance in hemodialysis patients.

Complement-mediated immune complex (IC) clearance was examined in the sera from regular hemodialysis patients, who accumulated excess factor D in the circulation. Both complement activities for the inhibition of insoluble IC formation and the solubilization of insoluble IC were lower in the sera from these patients than in the sera from normal individuals. A similar decrease in complement activities was observed in normal serum fortified with excess factor D. In addition, complement-processed IC was found to be bound less effectively to human erythrocytes via complement receptor type 1 in the sera from these patients and from normal serum fortified with excess factor D than in normal serum. Gel-filtration HPLC analysis revealed a decrease in covalent binding of C3 fragments to IC solubilized in the sera from hemodialysis patients or in normal serum fortified with excess factor D. This supports the notion that lower complement-mediated IC clearance is due to the paucity of complement fragments bound to IC. It is suggested that some detrimental alterations in the complement system, probably due to excess factor D, cause decreased IC clearance activity in hemodialysis patients with chronic renal failure.

Antigen-Antibody Complex↗
PubMed1993

Early branchings in the evolution of eukaryotes: ancient divergence of entamoeba that lacks mitochondria revealed by protein sequence data.

Phylogenetic analyses of ribosomal RNA sequences have played an important role in the study of early evolution of life. However, Loomis and Smith suggested that the ribosomal RNA tree is sometimes misleading--especially when G+C content differs widely among lineages--and that a protein tree from amino acid sequences may be more reliable. In this study, we analyzed amino acid sequence data of elongation factor-1 alpha by a maximum likelihood method to clarify branching orders in the early evolution of eukaryotes. Contrary to Sogin et al.'s tree of small-subunit ribosomal RNA, a protozoan species, Entamoeba histolytica, that lacks mitochondria was shown to have diverged from the line leading to eukaryotes with mitochondria before the latter separated into several kingdoms. This indicates that Entamoeba is a living relic of the earliest phase of eukaryotic evolution before the symbiosis of protomitochondria occurred. Furthermore, this suggests that, among eukaryotic kingdoms with mitochondria, Fungi is the closest relative of Animalia, and that a cellular slime mold, Dictyostelium discoideum, had not diverged from the line leading to Plantae-Fungi-Animalia before these three kingdoms separated.

Amino Acid Sequence↗
PubMed1993

Spontaneous rupture of splenic angiosarcoma: a case report of chemotherapeutic approach and review of the literature.

Angiosarcoma of the spleen is a rare disease, and the prognosis of this disease is extremely unfavorable. We herein review the case of a 45-year-old Japanese woman, who received a combined chemotherapy with cyclophosphamide, Adriamycin, vincristine, and prednisone after splenectomy and experienced a good response. The various types of chemotherapy for this disease are also discussed with references to the above case because no effective chemotherapeutic protocol for angiosarcomas has yet to be established.

Antineoplastic Combined Chemotherapy Protocols↗
PubMed1993

Primary structure and functional expression of the omega-conotoxin-sensitive N-type calcium channel from rabbit brain.

The complete amino acid sequence of a rabbit brain calcium channel (BIII) has been deduced by cloning and sequencing the cDNA. The open reading frame encodes 2339 amino acids, which corresponds to an M(r) of 261,167. A phylogenetic tree representing evolutionary relationships indicates that BIII is grouped together with the other rabbit brain calcium channels, BI and BII, into a subfamily that is distinct from the dihydropyridine-sensitive L-type subfamily. Transient expression in cultured skeletal muscle myotubes derived from muscular dysgenic mice demonstrates that the BIII channel mediates an omega-conotoxin-sensitive calcium current with kinetics and voltage dependence like those previously reported for whole-cell N-type current. Cell-attached patch recordings, with isotonic barium as the charge carrier, revealed distinct single channels with an average slope conductance of 14.3 pS.

Amino Acid Sequence↗