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Biomedical subjects

T Miyata

Publications and source records attributed to T Miyata.

At least 541 records · Page 30Linked to original sources
PubMed1989

A new surgical approach for treating infected epidermoid cysts using delayed primary closure.

A new surgical approach for treating infected epidermoid cysts was designed. This technique involves the lesion being incised and drained on the first day, 5-7 days after which it is removed together with the cyst wall excised parallel to Langer's tension lines. The wound is then closed by delayed primary closure. We employed this method in the treatment of 12 patients and observed the average time required for recovery (n = 12) was 18.6 +/- 2.5 days (mean +/- SD) and the number of days spent at the outpatient clinic, 10.2 +/- 2.6. There has been no recurrence or secondary infections in any of the patients to date.

Abscess↗
PubMed1989

Distribution of free N-acetylneuraminic acid in rat organs.

The concentration of free N-acetylneuraminic acid in various rat organs was estimated by gas chromatography/mass spectrometry. Its concentration was in the range of 3.95 to 104.72 micrograms/g wet tissues, being particularly high in the endocrine glands. The ratio of free N-acetylneuraminic acid to total N-acetylneuraminic acid varied from 0.031 to 0.183, being especially high in the adrenal gland (0.181) and heart (0.183).

Animals↗
PubMed1989

Newcastle disease virus evolution. I. Multiple lineages defined by sequence variability of the hemagglutinin-neuraminidase gene.

We compared the hemagglutinin-neuraminidase gene sequence among 13 strains of Newcastle disease virus (NDV) isolated over the last 50 years. Although overall homology was remarkably high, the sequence variability demonstrated the existence of at least three distinct lineages, which must have co-circulated for considerable periods. The sequence variability also appears to reflect some accumulation of mutations over time. Strictly correlating with the lineages, the translation products could be classified into three size classes. One class lacked the interchain disulfide bond, and another represented unusual precursor protein of biologically inactive form. The lineages correlated to some extent with virulence and place of isolation of the strains. However, antigenic variations, which were neither cumulative nor progressive, did not correlate with the lineages. These analyses showing multiple lineages were greatly facilitated by a precise calculation of synonymous substitutions, which had been largely free from selective pressures and had occurred frequently and evenly throughout the coding region.

Amino Acid Sequence↗
PubMed1989

Newcastle disease virus evolution. II. Lack of gene recombination in generating virulent and avirulent strains.

Sequence analysis and comparison of the fusion glycoprotein genes of 11 Newcastle disease virus (NDV) isolates indicated a high degree of functional and structural constraint exerted on the change of the glycoprotein. However, synonymous nucleotide substitutions occurred frequently throughout the coding region. Facilitated by an analysis of synonymous difference (Ks) in pairwise strain comparison, we defined the branching orders of the strains and identified three distinct evolutionary lineages correlating with the virulence as expressed by mean death time (MDT) for chick embryo. The typically virulent strains with MDT of about 50 hr were associated with one lineage, while the typically nonvirulent strains with MDT of infinity were of another lineage. The third lineage consisted of both virulent and avirulent strains whose MDTs lay on a continuum from 50 to 120 hr. Synonymous substitutions were found to occur with almost the same rates in the adjacent hemagglutinin-neuraminidase and membrane protein genes as in the fusion protein gene, and the branching orders based upon the Ks for these genes were essentially identical to those derived from the fusion protein gene. Therefore, no gene exchange by recombination seems to have occurred to generate the strains of distinct lineages. Rather, the different strains appear to have evolved through various degrees of accumulation of point mutations. Besides these evolutionary features, the present study strongly supports the importance of the previously identified signals for gene expression and for the proteolytic activation of the gene product.

Animals↗
PubMed1989

A clinicopathologic study of aneurysm formation of glutaraldehyde-tanned human umbilical vein grafts.

To clarify the process of graft degeneration and aneurysm formation we reviewed the angiographic and pathologic findings of three cases where aneurysms developed in glutaraldehyde-tanned human umbilical vein grafts. Seven aneurysms were detected in these three grafts 4 years after implantation. Six aneurysms originated from the bodies of the grafts and one originated from the factory-created anastomosis required for fabrication. Wrinkling and segmentation of the dilated graft documented by arteriography corresponded to mural dissection of the graft wall. Large protrusions near the aneurysms were revealed to be the transmurally dissected graft walls. At the site of dissection, blood escaped through the surrounding mesh of the graft to form a preaneurysmal change. Two aneurysms developed without breakdown of the outer mesh. The aneurysm of factory-created sutures was caused by tearing of the graft wall. Degeneration of human umbilical vein grafts, which appears to ensue inevitably with the passage of time after implantation, leads to the tear of the graft wall and transmural dissection to form aneurysm. In our department, candidates for human umbilical vein graft implantation have recently been restricted to the patients whose life expectancy is limited to only a few years because of the risk of aneurysm formation.

Aged↗
PubMed1989

Coagulation factor XII (Hageman factor) Washington D.C.: inactive factor XIIa results from Cys-571----Ser substitution.

Structural studies on a congenital abnormal coagulation factor XII (Hageman factor), factor XII Washington D.C., have been performed to identify the defect responsible for its lack of procoagulant activity. Amino acid sequence analysis of a tryptic peptide isolated from the abnormal factor XII indicated that Cys-571 (equivalent to Cys-220 in the chymotrypsin numbering system) had been replaced by serine. No other substitutions in the active-site triad--namely, His-393, Asp-442, and Ser-544--were found. We propose that the Cys-571----Ser replacement found in this factor XII variant destroys the formation of the disulfide linkage between Cys-540 and Cys-571, giving rise to an altered conformation of the active-site serine residue or the secondary substrate-binding site and, thus, leads to the loss of enzyme activity.

Amino Acid Sequence↗
PubMed1989

Evolutionary relationship of archaebacteria, eubacteria, and eukaryotes inferred from phylogenetic trees of duplicated genes.

All extant organisms are though to be classified into three primary kingdoms, eubacteria, eukaryotes, and archaebacteria. The molecular evolutionary studies on the origin and evolution of archaebacteria to date have been carried out by inferring a molecular phylogenetic tree of the primary kingdoms based on comparison of a single molecule from a variety of extant species. From such comparison, it was not possible to derive the exact evolutionary relationship among the primary kingdoms, because the root of the tree could not be determined uniquely. To overcome this difficulty, we compared a pair of duplicated genes, elongation factors Tu and G, and the alpha and beta subunits of ATPase, which are thought to have diverged by gene duplication before divergence of the primary kingdoms. Using each protein pair, we inferred a composite phylogenetic tree with two clusters corresponding to different proteins, from which the evolutionary relationship of the primary kingdoms is determined uniquely. The inferred composite trees reveal that archaebacteria are more closely related to eukaryotes than to eubacteria for all the cases. By bootstrap resamplings, this relationship is reproduced with probabilities of 0.96, 0.79, 1.0, and 1.0 for elongation factors Tu and G and for ATPase subunits alpha and beta, respectively. There are also several lines of evidence for the close sequence similarity between archaebacteria and eukaryotes. Thus we propose that this tree topology represents the general evolutionary relationship among the three primary kingdoms.

Adenosine Triphosphatases↗
PubMed1989

Fibrinogen Nagoya, a replacement of glutamine-329 by arginine in the gamma-chain that impairs the polymerization of fibrin monomer.

Structural studies on a hereditary abnormal fibrinogen, fibrinogen Nagoya (Takamatsu, J., Ogata, K., Kamiya, T., Koie, K., Takagi, T., & Iwanaga, S. (1979) Thromb. Haemost. 42, 78), were performed to identify the abnormality responsible for the impaired polymerization of fibrin monomer. Upon sodium dodecyl sulfate-polyacrylamide gel electrophoresis under reducing conditions, fibrinogen Nagoya showed the presence of an extra protein band with an apparent molecular weight of 49,500 in addition to the normal three subunit chains. Amino acid sequence analysis of a peptide isolated from a lysyl endopeptidase digest of one of the CNBr fragments derived from fibrinogen Nagoya indicated that Gln-329 in the gamma-chain had been replaced by Arg. This substitution can be explained by a single nucleotide change in the codon for Gln-329 (CAG----CGG). We conclude that Gln-329 in the gamma-chain is indispensable for the normal polymerization of fibrin monomer.

Amino Acid Sequence↗
PubMed1989

The NH2-terminal structures of human and rat liver microsomal NADH-cytochrome b5 reductases.

Detergent-solubilized NADH-cytochrome b5 reductase was purified from human liver microsomes. Both the purified enzyme and the membrane-binding domain isolated from the purified enzyme were determined to be modified at the NH2-terminal amino acid, glycine, by myristic acid in an amide form. Myristic acid was identified as a methyl ester by gas chromatography after the acid methanolysis of the purified enzyme and the NH2-terminal peptide. The NH2-terminal structure of the membrane-binding domain was determined to be CH3(CH2)12-CO-Gly-Ala-Gln-Leu-Ser-Thr-Leu-Gly-His-Met-Val-Leu-Phe-Pro-Va l- Trp-Phe-Leu-Tyr-Ser-Leu-Leu-Met-Lys. The sequence from Leu-7 to Lys-24 completely coincided with that deduced from the base sequence of complementary DNA (cDNA) from human placenta (Yubisui, T. et al. (1987) Proc. Natl. Acad. Sci. U.S. 84, 3609-3613). The NH2-terminal structure of the detergent-solubilized enzyme from rat liver microsomes was also analyzed for comparison with that of human liver microsomal enzyme. The NH2-terminal myristic acid and the first 7 amino acids of the membrane-binding domains of human, rat, and steer liver microsomal enzymes are completely conserved, and more than 70% homology was observed over the whole membrane-binding domains, implying the importance of the conserved structure as an anchor of the enzyme to the membrane.

Amino Acid Sequence↗
PubMed1989

Overexpression and purification of the recombinant Ca2+-binding protein, apoaequorin.

The small, monomeric Ca2+-binding photoprotein, aequorin, emits blue light by an intramolecular reaction when mixed with Ca2+. The photoprotein is made up of coelenterazine and molecular oxygen, bound noncovalently to apoaequorin (apoprotein). The chemical steps leading to light emission, involving the oxidative degradation of coelenterazine, have been studied extensively, but little is known about the active site and how the molecule catalyzes the oxidation of coelenterazine. The three-dimensional structure of the protein has not been determined and therefore answers to these questions have remained unavailable. The present paper describes a procedure for preparing fairly large amounts of apoaequorin and aequorin for X-ray crystallographic studies. It consists of fusing the apoaequorin cDNA to the signal peptide coding sequence of the outer membrane protein A of Escherichia coli, which is under the control of the lipoprotein promoter. When the cDNA was expressed in E. coli, a large excess of the recombinant protein was produced and released into the culture medium. Purification of the protein was accomplished by acid precipitation and DEAE-cellulose chromatography. The procedure yielded 7.4 mg of recombinant apoaequorin with a purity greater than 95% from 200 ml of culture medium. On regeneration with coelenterazine, the recombinant aequorin was fully active with Ca2+.

Aequorin↗
PubMed1989

Blood clotting factor IX Kashihara: amino acid substitution of valine-182 by phenylalanine.

Hemophilia B Kashihara is a severe hemorrhagic disorder in which the factor IX antigen is present in normal amounts but factor IX biological activity is markedly reduced. In addition, purified factor IX Kashihara is not activated by purified factor XIa in the presence of calcium ions. Amino acid sequence analysis of one of the tryptic peptides isolated from factor IX Kashihara indicated that Val-182 (equivalent to Val-17 in the chymotrypsin numbering system) had been replaced by Phe. No substitution was found in the members of the catalytic triad His-221, Asp-269, and Ser-365 of factor IX Kashihara. The Val-to-Phe replacement found in factor IX Kashihara appears to sterically hinder the cleavage of Arg 180-Val 181 by factor XIa required for the activation of this zymogen.

Amino Acids↗
PubMed1989

Primary structure of hemorrhagic protein, HR2a, isolated from the venom of Trimeresurus flavoviridis.

The complete amino acid sequence and disulfide bridge location of HR2a, one of the hemorrhagic proteins isolated from the snake venom of Trimeresurus flavoviridis, have been determined by analysis of peptides derived from digests with cyanogen bromide, lysyl endopeptidase, trypsin, and Staphylococcus aureus V8 protease. Peptides were purified by gel filtration followed by reversed-phase HPLC. HR2a has the amino-terminal sequence of less than Glu-Gln-Arg- and consists of a total of 202 residues with a calculated molecular weight of 23,015. Sequence analysis indicates the presence of another isoform which lacks the amino-terminal residue, making 201 amino acid residues with a molecular weight of 22,887. Three disulfide bridges of HR2a link Cys-118 to Cys-197, Cys-159 to Cys-181, and Cys-161 to Cys-164. HR2a contains a segment which is similar to the zinc-chelating sequences found in thermolysin and several mammalian metalloproteinases, suggesting that HR2a is a metalloproteinase with limited substrate specificity. However, there is no other significant sequence homology with thermolysin except for the zinc-ligand region.

Amino Acid Sequence↗
PubMed1989

Primary structure of H2-proteinase, a non-hemorrhagic metalloproteinase, isolated from the venom of the habu snake, Trimeresurus flavoviridis.

The complete amino acid sequence of and the locations of disulfide bridges in H2-proteinase, a major non-hemorrhagic proteinase isolated from the venom of the habu Trimeresurus flavoviridis, have been determined and compared with those of HR2a, one of the hemorrhagic metalloproteinases in this venom. The strategy involved consisted of structural analysis of peptides in digests with cyanogen bromide, lysyl endopeptidase, trypsin, Staphylococcus aureus V8 protease and thermolysin. Peptides were purified by gel filtration followed by reversed-phase HPLC. H2-proteinase is a non-glycosylated single chain polypeptide consisting of 201 amino acids with an amino-terminal pyroglutamic acid, a calculated molecular weight of 22,991 and a net charge of +14 at neutral pH. There was no evidence of heterogeneity of the sequence. H2-proteinase has a typical zinc-chelating sequence and its overall sequence identity with HR2a is 73.6%. The 3 disulfide bridges in H2-proteinase link Cys-117 to Cys-196, Cys-158 to Cys-180, and Cys-160 to Cys-163, in the same manner as in the case of HR2a. In striking contrast to HR2a, it contains en extra free cysteine residue at position 94 which becomes reactive to a sulfhydryl reagent in the presence of a denaturant.

Amino Acid Sequence↗
PubMed1989

Antimicrobial peptides, isolated from horseshoe crab hemocytes, tachyplesin II, and polyphemusins I and II: chemical structures and biological activity.

Tachyplesin is an antimicrobial peptide recently found in the acid extract of hemocytes from the Japanese horseshoe crab (Tachypleus tridentatus) [Nakamura, T. et al. (1988) J. Biol. Chem. 263, 16709-16713]. In our continuing studies on the peptide, we have found an isopeptide, tachyplesin II, and also polyphemusins I and II in hemocytes of the American horseshoe crab (Limulus polyphemus). The complete primary structures of these peptides, which are very similar to that of the previously isolated peptide, now named tachyplesin I, were determined to be as follows: (Table: see text). The isopeptide, tachyplesin II, consists of 17 residues with a COOH-terminal arginine alpha-amide. On the other hand, both polyphemusins I and II were found to contain 18 residues due to an additional Arg residue at the NH2-terminal end as well as a COOH-terminal arginine alpha-amide. The disulfide linkages for polyphemusin I consisted of two bridges between Cys-4 and Cys-17 and between Cys-8 and Cys-13, which was identical to in the case of tachyplesin I. Moreover, all of these peptides inhibited the growth of not only Gram-negative and -positive bacteria but also fungi, such as Candida albicans M9. Furthermore, complex formation between these peptides and bacterial lipopolysaccharides was also observed in a double diffusion test. These results suggest that tachyplesins and polyphemusins are probably located in the hemocyte membrane, where they act on antimicrobial peptides as a self-defense mechanism in the horseshoe crab against invading microorganisms.

Alkylation↗
PubMed1989

Factor IX Kawachinagano: impaired function of the Gla-domain caused by attached propeptide region due to substitution of arginine by glutamine at position -4.

Factor IX Kawachinagano (KWC) is a mutant factor IX protein initially recognized in a patient with severe haemophilia B, who had 46% of normal factor IX antigen and no detectable clotting activity. Previous studies indicated that factor IX KWC was not activated by factor XIa in the presence of Ca ions. In the present study, we purified and analysed factor IX KWC at a structural level in an attempt to clarify the nature of the impaired reaction with factor XIa. Kinetic studies showed that activation of factor IX KWC by factor X activator from Russell's viper venom (RVV-X) was normal, whereas activation by factor XIa was defective. Amino acid sequence analysis of tryptic peptides and direct analysis of the NH2-terminal sequence of factor IX KWC demonstrated that this mutant factor IX retained the propeptide region of 18 amino acids due to a substitution of arginine-(-4) by glutamine. These data suggested that the Gla-domain of factor IX KWC was dysfunctional, although the total gamma-Gla content, measured by alkaline-hydrolysis, was normal. We assumed that this attached propeptide region of the molecule directly interferes with the adjacent NH2-terminus and prevents the metal-induced conformational changes which are essential for biological activity of normal factor IX.

1-Carboxyglutamic Acid↗
PubMed1989

Epoxy compounds as a new cross-linking agent for porcine aortic leaflets: subcutaneous implant studies in rats.

The epoxy group of the epoxy compounds has an oxygen arm that can work as a flexible joint in a cross-linking bridge and can block not only the amino group but also the carboxy group of collagen peptide. The purpose of this study is to evaluate the anticalcification efficacy of the epoxy compounds as a cross-linking agent for xenograft bioprostheses. Porcine aortic leaflets were treated with 2% epoxy compounds and implanted in subcutaneous layer of 4-week-old rats. Measurement of calcium content showed that epoxy-treated implants received a minimal calcification: mean 0.64 micrograms/mg dry weight leaflet tissue (range 0.5-0.8; N = 7) at 1 month; mean 0.94 microgram/mg (range 0.3-1.3; N = 9) at 2 months; and mean 1.2 micrograms/mg (range 0.5-2.1; N = 10) at 3 months. Natural leaflets contained calcium of mean 0.43 microgram/mg. By contrast, glutaraldehyde-preserved implants were severely calcified: mean 91 micrograms/mg (range 41-130; N = 11) at 1 month; mean 136 micrograms/mg (range 73-205; N = 16) at 2 months; and mean 170 micrograms/mg (range 90-214; N = 21) at 3 months. The epoxy compounds provide more pronounced anticalcification effects than the glutaraldehyde under pressure load-free subcutaneous circumstances.

Animals↗
PubMed1989

[A strain of Pseudomonas vesicularis isolated from shower hose which supports the multiplication of Legionella].

In Japan, a fatal case due to Legionella micdadei was first recognized in our laboratory in 1986. On the epidemiological study just after the case, no Legionella was detected from the environmental samples of the patient's residence, such as shower water, tank water and so on. In the course of prospective investigations, no Legionella was isolated, but many organisms were grown on BCYE alpha and MWY agar plates. In the retrospective study, one of these organisms was found to support satellite growth of Legionella on BCYEagar without L-cysteine. This was the isolate from the shower hose and identified as Pseudomonas vesicularis with the biochemical and DNA-DNA hybridization test. And P. vesicularis type strain ATCC11426 also supported satellite growth of Legionella. Especially in the water supply system, the existence of P. vesicularis seemed to be effective on the growth of Legionella. It must be taken into consideration that efforts made to isolate the nutrient produced organisms as well as Legionella are needed.

Legionella↗