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Biomedical subjects

M Yamaguchi

Publications and source records attributed to M Yamaguchi.

At least 1,549 records · Page 86Linked to original sources
PubMed1985

Mitochondrial nicotinamide nucleotide transhydrogenase: inhibition by ethoxyformic anhydride, dansyl chloride, and pyridoxal phosphate.

Mitochondrial energy-linked nicotinamide nucleotide transhydrogenase (TH; EC 1.6.1.1) was inactivated by treatment with pyridoxal phosphate, ethoxyformic anhydride (EFA) or dansyl chloride. NADP and NADPH, but not NAD and NADH, protected TH against inhibition by pyridoxal phosphate, and L-lysine reversed this inhibition. The results suggested modification of an essential lysyl residue by pyridoxal phosphate, possibly at the NADP(H) binding site of TH. EFA and dansyl chloride inhibited TH in a similar manner. The effect of pH on the rate of inhibition of TH by EFA and dansyl chloride was the same, and in both cases addition of NADP and particularly NADPH accelerated the rate of inhibition, while addition of NAD or NADH had no effect. Double inhibition studies, using in one experiment dithiothreitol-reversible inhibition by 5,5'-dithiobis(2-nitrobenzoic acid) to protect the thiol groups of TH, and in another experiment lysine-reversible inhibition by pyridoxal phosphate to protect the putative essential lysyl residues of the enzyme, followed in each case by further treatment of the protected TH with EFA or dansyl chloride, suggested that the inhibitions by EFA and dansyl chloride were independent of the inhibitions by 5,5'-dithiobis (2-nitrobenzoic acid) and pyridoxal phosphate. The inhibitors discussed above are interesting, because pyridoxal phosphate is the only reagent known which appears to modify an essential residue in the NADP(H), but not the NAD(H), binding site of TH, and EFA and dansyl chloride are the only inhibitors known which appear to react with essential residues outside the active site of TH. It is possible that EFA and dansyl chloride inhibitions involve modification of essential prototropic residues in the proton translocation domain of the enzyme.

Animals↗
PubMed1985

Determination of alpha-keto acids in serum and urine by high-performance liquid chromatography with fluorescence detection.

A simple, rapid and highly sensitive high-performance liquid chromatographic method for the determination of alpha-keto acids in serum and urine is described. In dilute hydrochloric acid, alpha-keto acids are converted by 1,2-diamino-4,5-dimethoxybenzene into highly fluorescent quinoxalinone derivatives. The derivatives are isocratically separated simultaneously within 14 min by reversed-phase chromatography on a Radial-Pak cartridge C18 and detected fluorimetrically. The limits of detection are 10-300 fmol in an injection volume of 10 microliters (40-1200 pmol/ml of serum or urine). This sensitivity permits precise determination of several alpha-keto acids in 5 microliter of serum or urine from healthy persons, and also the determination of phenylpyruvic acid in normal urine which cannot be simultaneously determined by other methods.

Adult↗
PubMed1985

Photoaffinity labeling of D-(-)-beta-hydroxybutyrate dehydrogenase by (arylazido)-beta-alanyl-substituted nicotinamide adenine dinucleotide.

(Arylazido)-beta-alanyl-substituted nicotinamide adenine dinucleotide (N3-NAD) is a photosensitive analogue of NAD capable of photoinduced nitrene generation and insertion into a nearby molecule. In the dark, N3-NAD can replace NAD as a cosubstrate for the mitochondrial D-(-)-beta-hydroxybutyrate dehydrogenase (BDH). With purified, phospholipid-reconstituted BDH and NAD as the variable substrate, the apparent Km and Vmax values were respectively 0.25 mM and 62.5 mumol min-1 (mg of protein)-1. With N3-NAD as the variable substrate, these values were respectively 0.59 mM and 5 mumol min-1 (mg of protein)-1. Photoirradiation of BDH in the presence of N3-NAD resulted in irreversible inhibition of the enzyme and incorporation into the protein of radioactivity from tritiated N3-NAD. Photoirradiation of BDH plus or minus NAD in the absence or presence of (arylazido)-beta-alanine caused little or no inhibition. The photoinhibition of BDH in the presence of N3-NAD was prevented nearly completely by addition of NADH, NAD plus beta-hydroxybutyrate, or NAD plus 2-methylmalonate and partially by addition of NAD. Moreover, the presence of NADH prevented, and prior partial modification of BDH at the NAD(H)-protectable site by N-ethylmaleimide decreased, the incorporation of radioactivity into BDH from photoirradiated [3H]N3-NAD. The above results suggest that N3-NAD can be used for photoaffinity labeling of BDH at the active site.

Affinity Labels↗
PubMed1985

Fine structure of wide and narrow vertebrate muscle Z-lines. A proposed model and computer simulation of Z-line architecture.

A model of the structure of vertebrate Z-lines and Z-line analogs is introduced and supported by evidence from electron microscope studies of wide Z-lines (rat and feline soleus, and feline and canine cardiac muscles), narrow Z-lines (guppy, newt and frog skeletal muscles), and Z-rods (from a patient with nemaline myopathy and from cardiac muscles of aged dog). The model is based on a pair of Z-filaments (termed a Z-unit), which are linked near their centers at a 90 degrees angle and form bridges between neighboring antipolar thin (actin) filaments. A square lattice of four Z-filament pairs (the basic structure of the Z-line, termed a Z-line unit) defines the geometrical position of the I-square unit. In this native state of the Z-line, small square and large square net forms appear in cross-section. Other cross-sectional patterns of Z-lines, including basket-weave and diagonal-square net patterns, can be explained by detachment of the Z-filament from the Z-filament binding region within each Z-filament pair due to chemical or physical stress. Dissection of Z-lines and Z-line analogs with calcium-activated neutral protease provides evidence that the width of all wide Z-line structures is determined by the amount of overlap of antipolar thin filaments from adjacent sarcomeres. Longitudinal patterns of narrow and wide Z-lines are shown and described in relation to the model. To test the proposed model, the dynamics of the Z-line unit structure were computer-simulated. An attempt was made to correlate longitudinal (z direction) and cross-sectional (x and y directions) patterns and to determine the amount of movement of thin or Z-filaments that is required to explain the diversity observed in cross-sectional patterns of Z-lines. The computer simulations demonstrated that the structural transitions among the small square, and therefore large square net, as well as basket-weave and diagonal-square net forms seen in cross-sections could be caused by movements of thin filaments less than 10 nm in any direction (x, y or z).(ABSTRACT TRUNCATED AT 400 WORDS)

Actins↗
PubMed1985

Effect of ibuprofen on the healing phase of experimental myocardial infarction in the rat.

The healing phase of acute myocardial infarction (AMI) is initiated by proteolysis of necrotic myocardium, followed by infiltration of fibroblasts and deposition of collagen. To assess whether ibuprofen, a potent antiinflammatory agent, preserves existing collagen and enhances deposition of new collagen during infarct healing, biochemical and morphologic studies were made of experimentally induced myocardial infarcts in untreated rats and in rats treated with ibuprofen. All treated rats received 12.5 mg/kg of ibuprofen at 1, 6 and 18 hours after AMI. Group 1 rats underwent measurement of myocardial hydroxyproline (HP) content at 24 hours after AMI. Group 2 rats received ibuprofen, 12.5 mg/kg, twice a day for 2 additional days, with measurement of myocardial HP at 3 days (group 2a) or 21 days (group 2b) after AMI. Group 3 rats received ibuprofen, 12.5 mg/kg, twice a day for 6 additional days with measurement of HP content, or infarct size and degree of thinning at 21 days after AMI. Compared with untreated rats, ibuprofen-treated rats had significantly greater amounts of HP in the infarct at 24 hours (group 1, 8.9 +/- 2.2 nmol/mg dry weight vs untreated, 7.1 +/- 2.8 nmol/mg dry weight, p less than 0.04) and at 21 days (group 2b, 112 +/- 37 nmol/mg dry weight vs untreated, 91 +/- 39 nmol/mg dry weight, p less than 0.05, and group 3, 125 +/- 51 nmol/mg dry weight vs untreated, 91 +/- 39 nmol/mg dry weight, p less than 0.003). Substantial scar thinning was noted in all rats; no difference in scar thinning was noted between treated and untreated rats at 21 days after AMI.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1985

DNA primase-DNA polymerase alpha from simian cells. Modulation of RNA primer synthesis by ribonucleoside triphosphates.

DNA primase-DNA polymerase alpha, purified 53,000-fold from CV-1 cells, synthesized predominantly (p)ppA(pA)6-primed DNA on a poly(dT) template. About 80% of the RNA primers synthesized on an M13 DNA template were (p)ppA/G(pN)5-7, and 20% were (p)ppA/G(pN)0-4. RNA primer size was determined by gel electrophoresis after removing nascent DNA with phage T4 DNA polymerase 3'-5' exonuclease, leaving a single dNMP at the 3'-end of the RNA primer, and the terminal 5'-(p)ppN residue was determined by "capping" with [alpha-32P]GTP using vaccinia guanylyl-transferase. The processivity of DNA synthesis initiated by de novo synthesis of RNA primers was the same as that initiated on pre-existing RNA primers (10-15 dNMPs), although initiation on pre-existing primers was strongly preferred. Primers always began with A or G, even at high levels of CTP or UTP, although the ratio of A to G varied from 4:1 to 1:1 depending on the relative concentrations of ATP and GTP in the assay. ATP and GTP had no effect on primer length, but the fraction of shorter RNA primers increased 2-fold with higher concentrations of CTP or UTP. Nearest-neighbor analysis revealed a preference for purine ribonucleotides at RNA covalently linked to the 5'-end of DNA (RNA-p-DNA) junctions, and increasing the concentration of a single rNTP increased slightly its presence at RNA-p-DNA junctions. Thus, the base composition and size of RNA primers synthesized by DNA primase-DNA polymerase alpha is modulated by the relative concentrations of ribonucleoside triphosphates.

Animals↗
PubMed1985

Coincidence of H+ binding and Ca2+ dissociation in the sarcoplasmic reticulum Ca-ATPase during ATP hydrolysis.

H+ and Ca2+ concentration changes in the reaction medium following MgATP addition at pH 6.0 were determined with the partially purified Ca-ATPase from sarcoplasmic reticulum vesicles in the presence of 25-50 microM CaCl2 and 5 mM MgCl2 at 4 degrees C. Previously, we showed a sequential occurrence of H+ binding and H+ dissociation in the Ca-ATPase during ATP hydrolysis and further suggested that the H+ binding takes place inside the vesicles (Yamaguchi, M., and Kanazawa, T. (1984) J. Biol. Chem. 259, 9526-9531). The present results demonstrate that the H+ binding occurred coincidently with Ca2+ dissociation from the enzyme upon conversion of the phosphoenzyme (EP) intermediate from the ADP-sensitive form to the ADP-insensitive form in the catalytic cycle of ATP hydrolysis. As KCl decreased in the medium, the extent of the H+ binding increased almost proportionately with the extent of either the Ca2+ dissociation or the accumulation of ADP-insensitive EP. Both the H+ binding and the Ca2+ dissociation were prevented by a modification of the specific SH group of the enzyme essential for the conversion of ADP-sensitive EP to ADP-insensitive EP. In the late stage of the reaction, H+ dissociation from the enzyme occurred coincidently with Ca2+ binding to the dephosphoenzyme which was formed by EP decomposition. These results are consistent with the possibility that the H+ ejection during the Ca2+ uptake with the intact vesicles previously shown by several investigators takes place through a Ca2+/H+ exchange directly mediated by the membrane-bound Ca-ATPase.

Adenosine Diphosphate↗
PubMed1985

6-Keto prostaglandin F1 alpha, thromboxane B2, and 13,14-dihydro-15-keto prostaglandin F concentrations of normotensive and preeclamptic patients during pregnancy, delivery, and the postpartum period.

Plasma 6-keto prostaglandin F1 alpha, 13,14-dihydro-15-keto prostaglandin F, and thromboxane B2 levels were measured in normotensive and preeclamptic patients during pregnancy and the postpartum period. From 30 to 40 weeks of gestation, 6-keto prostaglandin F1 alpha levels of preeclamptic patients were significantly lower than those of normotensive women; 13,14-dihydro-15-keto prostaglandin F and thromboxane B2 concentrations in preeclamptic patients did not significantly differ from those of the normotensive group. At delivery, 6-keto prostaglandin F1 alpha levels of maternal and umbilical venous plasma of preeclamptic women were also significantly lower than those of normotensive women. In the postpartum period these three prostanoids were not significantly different in normotensive women compared to preeclamptic women, with clinical preeclamptic symptoms soon disappearing in most of our patients. From the results, it seems that prostacyclin plays an important role in preeclampsia.

6-Ketoprostaglandin F1 alpha↗
PubMed1985

Activity of a novel carotenoid analog, 4,4'-(3,7,12,16-tetramethyl-1,3,5,7,9,11,13,15,17-octadecanonaen -1,18- diyl) bis(1-ethylpyridinium) dibromide (Y-18598) on mouse leukemia P388.

4,4'-(3,7,12,16-Tetramethyl-1,3,5,7,9,11,13,15,17-octadecanonaen-1 ,18-diyl) bis (1-ethylpyridinium) dibromide (Y-18598), a novel carotenoid analog with 1-ethylpyridine rings as terminal groups, was found to be effective against P388 leukemia which was refractory to amphotericin B and retinoic acid used as representatives of polyene-containing drugs. The life span of leukemic mice was increased by 56% over the control following administration of 2.5 mg/kg on days 1-5. Among these polyene-containing compounds, Y-18598 was the most highly cytotoxic to P388 leukemia cells in vitro.

Amphotericin B↗
PubMed1985

Polysaccharides sulfated at the time of gastrulation in embryos of the sea urchin Clypeaster japonicus.

Based on the fact that the development of sea urchin embryos is arrested at the blastula stage in sulfate-free sea water (SFSW), we attempted in the present study to elucidate the nature of sulfated polysaccharides (PSs) which appear at the time of gastrulation in embryos of the sea urchin Clypeaster japonicus. Electrophoretic analysis of PSs prepared from embryos at different developmental stages revealed that three kinds of PSs (3A, 3B, 3C) appear de novo at the gastrula stage, and that these PSs are not found in embryos at the hatching blastula stage, nor are they found in permanent blastula reared in SFSW. These, three PSs were mostly of extracellular matrix origin. Among them, 3C was identified as dermatan sulfate on the basis of its electrophoretic mobility and sensitivity to enzymatic digestion. 3A and 3B remained to be identified. Further, a plausible precursor of 3C, which was sulfated under normal conditions, was detected as 6D in the embryos reared in SFSW. Autoradiographic analysis using [35S]sulfate revealed that these three PSs, accounted for more than 90% of [35S]sulfate incorporated into the acid PS fraction during gastrulation.

Animals↗
PubMed1985

Stimulatory effect of calcitonin on fatty acid synthesis in the liver of fed rats.

The effect of calcitonin (CT) on free fatty acid concentration in the serum and liver of fed rats was investigated. A single subcutaneous administration of CT (synthetic [Asu1,7] eel CT;80 MRC mu/100 g body weight) produced a significant increase in serum free fatty acid concentration. An appreciable effect of CT was observed at a dose of 5 MRC mU/100 g body weight. The hormonal effect was also observed in thyroparathyroidectomized rats. The effect of CT on serum free fatty acid was diminished by fasting. Free fatty acid content in the hepatic cytosol of fed rats was markedly increased by CT administration. The hormonal effect was observed at a dose of 5 MRC mU/100 g body weight. Furthermore, stimulation of fatty acid synthesis caused by intraperitoneal injection of alanine (1.122 mmoles/100 g body weight) was markedly enhanced by administration of CT (5, 20 and 80 MRC mU/100 g body weight). This effect of CT on the liver may be the cause of increased level of fatty acid in the serum. The present results suggest that CT may stimulate synthesis of free fatty acid in the liver of fed rats.

Alanine↗
PubMed1985

Positional specificity of lysosomal acid lipase purified from rabbit liver.

The hydrolysis of tri-, di-, and monooleoylglycerols by highly purified lysosomal acid lipase from rabbit liver (Imanaka, T., et al. (1984) J. Biochem. 96, 1089-1101) was examined. The degradative products were separated by thin layer chromatography on silicic acid impregnated with boric acid. Trioleoylglycerol was hydrolyzed with intermediate accumulation of 1,2(2,3)-dioleoylglycerol and 2-monooleoylglycerol, but no detectable production of the 1,3-isomer. 1,2(2,3)-Dioleoylglycerol was also hydrolyzed with the intermediate accumulation of 2-monooleoylglycerol. The 1(3)-isomer of monooleoylglycerol was hydrolyzed exclusively, with no hydrolysis of the 2-isomer. These results indicate that the enzyme shows preference for 1(3)-ester bonds and that the main lipolytic reaction sequence catalyzed is triacylglycerol---1,2(2,3)-diacylglycerol---2-monoacylglycerol.

Animals↗