Search PubMed⌕ Search

Biomedical subjects

Y Miyake

Publications and source records attributed to Y Miyake.

At least 667 records · Page 37Linked to original sources
PubMed1980

Gas chromatography combined with mass spectrometry for the identification of organic sulfur compounds in shellfish and fish.

The authors determined that the organic sulfur compounds usually contained in crude oil can be used as a marker of oil pollution in shellfish and fish. Short-necked clams (Tapes Angdala phillipinarum A. et Reeve) and eels were maintained in a controlled laboratory environment in water with suspension of crude oil. Mass spectra and mass chromatograms of short-necked clam extract showed the presence of organic sulfur compounds of alkyl- (from C1 to C10) benzothiophenes, dibenzothiophene, alkyl- (C1 to C8) dibenzothiophene, naphthalene, and alkyl- (C1 to C6) naphthalene. Alkyl benzothiophenes and alkyl dibenzothiophenes of low carbon numbers were more easily transferred to short-necked clams than those of higher carbon numbers. Capillary column gas chromatography-mass chromatograms of crude oil and extract from the soft body of a short-necked clam showed the presence of organic sulfur compounds of alkyl benzothiophene (from C2-C6), dibenzothiophene, and alkyl dibenzothiophene (C1-C4). Besides sulfur components, various other compounds were contained in crude oil and short-necked clam, namely alkyl benzene (C3-C9), naphthalene, alkyl naphthalene (C1-C5), phenanthrene, alkyl phenanthrene (C1-C6), n-paraffin (C12-C21), and probably pyrene and alkyl pyrene (C1-C2). Also, mass chromatograms of crude oil and the extract from eel flesh showed the presence of alkyl benzothiophene (C1-C5), dibenzothiophene, naphthalene, and alkyl naphthalene (C1-C5). Data indicated that the organic sulfur compounds and polyaromatic compounds could serve as markers of oil pollution in shellfish and fish. for the identification of sulfur compounds by capillary gas chromatography/mass spectrometry, a new intensity matching method was devised.

Animals↗
PubMed1980

Studies on the reaction of D-amino acid oxidase with beta-cyano-D-alanine. Observation of an intermediary stable charge transfer complex.

The reaction of D-amino acid oxidase [EC 1.4.3.3] (DAO) from porcine kidney with beta-cyano-D-alanine (D-BCNA) was studied. DAO was found to catalyze elimination of the cyano group as well as oxidation of D-BCNA. During the course of the reaction in the presence of excess oxygen, an intermediate was observed which exhibited a characteristic absorption spectrum with a broad charge transfer band in the longer wavelength region. The CD spectrum of this intermediate resembles that of DAO-anthranilate complex. The rate of oxygen consumption in the aerobic reaction decreased with time, suggesting product inhibition due to complex formation between the enzyme and the product. Anaerobic addition of D-BCNA reduced the enzyme to its fully reduced state, the CD spectrum of which closely resembles that of the enzyme reduced by excess D-alanine. When an appropriate amount of D-BCNA was added to the enzyme under air, the charge transfer complex was observed immediately, and underwent a change to the reduced state as the oxygen was consumed. The binding strength in the charge transfer complex was found to be comparable to that in DAO-benzoate complex. The accumulating product in the oxidation of D-BCNA had a strong absorption at 285 nm. The aerobic reaction of beta-cyano-L-alanine (L-BCNA) with snake venom L-amino acid oxidase (LAO) produced the same product with an absorption at 285 nm as the reaction of DAO with D-BCNA. The product obtained in the reaction with LAO was found to form the same charge transfer complex with DAO. We tentatively identified this product as alpha-amino-beta-cyanoacrylate and the charge transfer complex as the complex of alpha-amino-alpha-cyanoacrylate with the oxidized enzyme. A hypothetical reaction pathway based on the present finding is proposed. Addition of L-BCNA to the enzyme produced an absorption spectrum very similar to that of the DAO-benzoate complex without oxidation or elimination. L-BCNA was found to be a competitive inhibitor of the oxidation of D-alanine.

Alanine↗
PubMed1980

Studies of maternal and fetal 11-deoxycorticosterone, corticosterone,18-hydroxycorticosterone and aldosterone in normal and toxemic pregnancy.

Simultaneous determination of maternal and fetal plasma 11-deoxycorticosterone (DOC), corticosterone (B), 18-hydroxycorticosterone (18OH-B) and aldosterone (ALDO) in normal and toxemic pregnancies was performed by utilizing a newly developed radioimmunoassay. The mean levels of plasma DOC and ALDO in normal term pregnancy were significantly higher than those of nonpregnant women. Comparing the mean levels of umbilical arterial plasma to those in maternal plasma at delivery, DOC and ALDO were consistently higher in the former, while B was higher in the latter, and 18OH-B was largely unchanged. However, no umbilical arteriovenous gradient was found in any corticosteroid measured. In toxemia of pregnancy, the corticosteroid levels in the maternal plasma were lower than those in normal pregnancies, and in umbilical arterial plasma, B was significantly higher and 18OH-B and ALDO remained almost the same as those in normal pregnancy. Thus, it appears that ALDO synthesis in mother in normal term pregnancy was enhanced but in toxemia of pregnancy it was suppressed, while in the fetus, it was enhanced not only in normal pregnancy but also in toxemia of pregnancy.

18-Hydroxycorticosterone↗
PubMed1980

Circular dichroism studies on flavoproteins containing covalently bound coenzymes.

Absorption and circular dichroism spectra of cholesterol oxidase from Schizophyllum commune and choline oxidase from Alcaligenes sp. were measured and compared. The prosthetic group of cholesterol oxidase is 8 alpha-[N(1)-histidyl]-FAD (1, 2), while that of choline oxidase is 8 alpha-[N(3)-histidyl]-FAD (3). In the CD spectra of the two enzymes in either the oxidized or reduced state, the corresponding bands in the visible region are of approximately the same intensity and shape but of opposite sign. A notable feature in the CD spectra of the two enzymes after light irradiation is the appearance of a CD band in the longer wavelength region (550-650 nm) and the opposite signs of the CD band in this region in the two enzymes. The similarity of the shape and intensity of the CD spectra of the two enzymes suggests that the environments surrounding the flavin moieties are very similar, and the sign reversal of the CD bands suggests that the mutual orientations between the transition moment of flavin and that of its environment differ in the two enzymes.

3-Hydroxysteroid Dehydrogenases↗
PubMed1980

Identification and properties of the prosthetic group of choline oxidase from Alcaligenes sp.

Choline oxidase from Alcaligenes sp. catalyzed the oxidation of choline and betaine aldehyde to betaine with concomitant consumption of oxygen and production of hydrogen peroxide. The values of Km for choline and betaine aldehyde were 0.87 and 6.2 mM, respectively. The molecular weight of the enzyme was estimated to be 66,000 by SDS-gel electrophoresis and 72,000 by gel-filtration using a high performance liquid chromatograph. The prosthetic group of the enzyme was identified as 8 alpha-[N(3)-histidyl]-FAD from the electrophoretic mobility at pH 6.25 of the hydrolysate of the methylated histidylflavin. The visible absorption spectrum of the enzyme showed peaks at 358 and 453 nm and a shoulder at about 480 nm. The covalently bound FAD was reduced on addition of either choline or betaine aldehyde under anaerobic conditions and was reoxidized by aeration. The enzyme was found to contain 1 mol of FAD per mol enzyme. Amino acid analysis of a purified flavin peptide gave the following molar ratios of amino acids to flavin: pro(1), Asp + Asn(3), Ser(1), His(1), and Arg(1). Aspartic acid was the N-terminal amino acid. The partial sequence of amino acids in the flavin peptide was as follows: Formula (See Text).

Alcaligenes↗
PubMed1979

Identification of the covalently bound flavin prosthetic group of cholesterol oxidase.

Highly purified preparations of cholesterol oxidase from Schizophyllum commune contain a covalently bound flavin component. A flavin peptide has been obtained by digestion with trypsin-chymotrypsin and purification on a column of phosphocellulose. Digestion with nucleotide pyrophosphatase results in increased fluorescence at pH 3.4 and release of 5'-adenylate, showing that the flavin is in the dinucleotide form. The absorption spectrum of the flavin peptide shows the hypsochromic shift of the second absorption band characteristic of 8 alpha-substituted flavins. The fluorescence at pH 7 is extensively quenched even in the mononucleotide form, with a pKa at pH 5.8 in the flavin peptide and at 5.05 following acid hydrolysis to the aminoacyl flavin level. This suggests that histidine is the amino acid substituted at the 8 alpha position of the flavin and that N(1) of the imidazole ring is the site of attachment. These data, the reduction of the flavin by borohydride, and comparison of the mobilities in high voltage electrophoresis at two pH values with N(1)- and N(3)-histidyl riboflavin and their 2',5'-anhydro forms shows that the prosthetic group of cholesterol oxidase is 8 alpha-[N(1)-histidyl]-FAD.

3-Hydroxysteroid Dehydrogenases↗
PubMed1979

Purification and characterization of a chymotrypsin-like enzyme (protease-S) in thermally injured rat skin.

Protease-S was extracted from thermally injured rat skin, and partially purified by column chromatography using Sephadex G-50, CM-Sephadex (A-50), Sephadex G-75 gel filtration. The optimum pH of this enzyme was 8.6--8.8, and the molecular weight determined by Sephadex G-75 gel filtration was approximately 30 000. This enzyme is active on the N-acetyl-L-tyrosine ethyl ester, N-succinyl-L-phenylalanine-p-nitroanilide (of chymotrypsin substrate) but not N-tosyl-L-arginine methyl ester, N-benzoyl-L-arginine-p-nitroanilide. Also, protease-S was completely inhibited by diisopropylfluorophosphate (1 mM) or phenylmethylsulfonylfluoride (10 micrometer), and N-tosyl-L-phenylalanine chloromethylketone (1 mM). These results are very similar to those obtained with bovine chymotrypsin. But the enzyme is not identical with the chymotrypsin-like proteases in mast cells and leukocyte granules. When proteases-S was measured during the inflammatory reaction in vivo, maximal activity was found after 8 h, at the end of inflammation.

Animals↗
PubMed1979

Reconstitution of lipid vesicles associated with HVJ (Sendai virus) sikes. Purification and some properties of vesicles containing nontoxic fragment A of diphtheria toxin.

A mixture of HVJ (Sendai virus) spike proteins, the nontoxic fragment A of diphtheria toxin, lecithin, and cholesterol was solubilized in sucrose solution containing a nonionic neutral detergent. The liposomal vesicles which formed on removal of the detergent by dialysis were purified by gel filtration and centrifugation on a sucrose gradient. The resulting purified vesicles had hemagglutinating activity, hemolytic activity and, after solubilization, the enzymic activity of fragment A. The vesicles had no cell fusion activity. Electron microscopy showed that both the outside and inside of membranes of the vesicles were associated with the spikes. When the vesicles were freeze-fractured, no large aggregates of particles were seen on either face. Such fragment A-containing lipid vesicles (liposomes) with HVJ spikes bound to mamalian cell membrane and released their fragment A into the cytoplasm causing cell death. Neither fragment A-containing liposomes without spikes nor empty liposomes with spikes were toxic.

Cell Fusion↗