Biomedical subjects

M Unno

Publications and source records attributed to M Unno.

At least 19 recordsLinked to original sources

PubMed1996

Role of Arg112 of cytochrome p450cam in the electron transfer from reduced putidaredoxin. Analyses with site-directed mutants.

The mechanism for the reduction of ferric cytochrome P450cam by reduced putidaredoxin, the physiological electron donor for the cytochrome, has been studied by using site-directed mutants of cytochrome P450cam, in which Arg112, an amino acid residue at the presumed binding site for putidaredoxin, was changed to several other amino acid residues. The affinity of reduced putidaredoxin for ferric cytochrome P450cam to form a diprotein complex was decreased greatly by changing Arg112 to a neutral amino acid such as Cys, Met, or Tyr. The rate of intracomplex electron transfer from putidaredoxin to cytochrome P450cam also diminished upon replacing the basic residue with neutral ones, being 42, 18, 4.0, 1.3, and 0. 16 s-1 for Arg (wild type), Lys, Cys, Met, and Tyr enzymes, respectively. Furthermore, the oxidation-reduction potential of cytochrome P450cam (Fe3+/Fe2+ couple) decreased in a similar way to the decrease in the rate of electron transfer upon amino acid substitution; the values were -138, -162, -182, -200, and -195 mV for Arg (wild type), Lys, Cys, Met, and Tyr enzymes, respectively. These results indicate that the amino acid substitution at position 112 affects the oxidation-reduction potential of the heme iron in cytochrome P450cam, thereby diminishing the rate of electron transfer between the two metal centers. The rate of electron transfer from putidaredoxin to oxyferrous cytochrome P450cam also diminished upon substitution of Arg112 with a neutral amino acid.

Arginine↗

PubMed1995

Nicotinamide inhibits IRF-1 mRNA induction and prevents IL-1 beta-induced nitric oxide synthase expression in pancreatic beta cells.

Nitric oxide produced by inducible nitric oxide synthase in islets exerts inhibitory and cytotoxic effects on pancreatic beta cells and is therefore thought to be a potent mediator in the pathogenesis of Type I diabetes mellitus. Here, using isolated rat pancreatic islets, we show that high-concentration nicotinamide (20 mM), but not low-concentration nicotinamide (5 mM), attenuates the interleukin-1 beta-evoked inhibition of glucose-induced insulin secretion by preventing the induction of interferon regulatory factor-1, a transcriptional factor which plays an essential role in inducible nitric oxide synthase gene expression, and the interleukin-1 beta-induced nitric oxide formation. High-concentration nicotinamide also restored an interleukin-1 beta-induced decrease in ATP content in pancreatic beta cells, suggesting that interleukin-1 beta-induced nitric oxide inhibits the mitochondrial function. The present results show the molecular basis of the preventive effect of high-dose nicotinamide on Type I diabetes mellitus.

Adenosine Triphosphate↗

PubMed1994

Characterization of the 5'-regulatory region of rat Reg I gene.

We report here the characterization of the 5'-regulatory region of rat Reg I gene encoding a growth stimulating factor for pancreatic beta-cells. Transient expression assays of the 5'-flanking region/luciferase fusion gene in AR4-2J cells showed that the -304/-237 region contained positive cis-acting elements. Gel shift assays using AR4-2J and rat pancreas nuclear extracts showed the formation of a specific complex with the -256/-237 oligonucleotide.

Animals↗

PubMed1994

High-pressure flash photolysis study of hemoprotein: effects of substrate analogues on the recombination of carbon monoxide to cytochrome P450CAM.

The effects of camphor and camphor analogues on the CO recombination kinetics of ferrous cytochrome P450CAM (P450CAM) at 293 K have been studied as a function of hydrostatic pressure (0.1-200 MPa) by means of flash photolysis. At 0.1 MPa, the association rate constant (kon) for substrate-free P450CAM is 8.5 x 10(6) M-1 s-1. Measurements as a function of pressure lead to a determination of the activation volume (delta V not equal to) of +4 cm3 mol-1 for substrate-free protein. This positive delta V not equal to is interesting because the CO association reaction of various hemoproteins, such as myoglobin and hemoglobin, exhibit negative delta V not equal to values [Adachi, S., & Morishima, I. (1989) J. Biol. Chem. 264, 18896-18901; Unno, M., Ishimori, K., & Morishima, I. (1990) Biochemistry 29, 10199-10205]. The binding of d-camphor and some camphor analogues (d-fenchone, 3-endo-bromocamphor, and 3,3,5,5-tetramethylcyclohexanone) into the heme pocket strongly influences the kinetics, i.e., kon is reduced ((1-10) x 10(5) M-1 s-1) and delta V not equal to is altered to a negative value (-14 to -32 cm3 mol-1). The negative delta V not equal to suggests that the effects of camphor and these camphor analogues are due to an increase in the iron-ligand bond formation barrier. On the other hand, the binding of adamantane and norcamphor does not affect the kinetics. This result is particularly surprising because both substrate analogues are located in the immediate vicinity of the CO binding site.(ABSTRACT TRUNCATED AT 250 WORDS)

Camphor↗

PubMed1994

Structure and expression of a novel rat RegIII gene.

We have isolated a rat cDNA and a novel gene, Reg (regeneration-promoting gene). The cDNA encodes a 174-amino-acid (aa) RegIII protein with a 25-aa signal peptide. The RegIII gene spans 2.7 kb and consists of six exons and five introns. RegIII was expressed in regenerating pancreatic islets, but not in normal islets.

Amino Acid Sequence↗

PubMed1994

Pancreatic beta-cell replication and amelioration of surgical diabetes by Reg protein.

We previously isolated from a rat regenerating islet cDNA library a gene named Reg, which is expressed in regenerating islets but is not expressed in normal islets. Here we examined the effect of rat Reg protein on pancreatic beta-cell replication using both 90% depancreatized rats and isolated islets. The depancreatized rats that received i.p. administration of recombinant rat Reg protein (1 mg/kg per day) for 2 months showed amelioration of the surgical diabetes, as evidenced by a significant decrease in blood glucose with an increased beta-cell mass in the residual pancreas. In isolated rat islets, Reg protein (18-180 nM: 0.3-3 micrograms/ml) significantly increased [3H]thymidine incorporation into the nuclei of beta cells. These results indicate that Reg protein is a growth factor for pancreatic beta cells and also suggest that the administration of Reg protein could be used as another therapeutic approach for diabetes mellitus.

Animals↗

PubMed1994

Isolation, structural determination and expression of a novel reg gene, human regI beta.

We have isolated a novel human gene and cDNA encoding a member of the regI proteins, regI beta. The gene encodes a 166-amino acid protein which has 22 amino acid substitutions in comparison with the previously isolated human reg protein, regI alpha. RegI beta was expressed only in pancreas, whereas regI alpha was expressed in kidney and stomach as well as in pancreas.

Amino Acid Sequence↗

PubMed1993

Effects of intra- and intersubunit hydrogen bonds on the R-T transition in human hemoglobin as studied with alpha 42(C7) and beta 145(HC2) mutations.

To clarify the effects of specific inter- and intrasubunit hydrogen bonds on the R-T transition in human hemoglobin (Hb A), the recombination reaction of carbon monoxide with artificial mutant Hbs was measured and analyzed. One of the hydrogen bonds we focused on is formed between Tyr-42 alpha and Asp-99 beta in the alpha 1-beta 2 interface of Hb A, which is one of the hydrogen bonds characteristic of the T state. Hb His-42 alpha, in which Tyr-42 alpha is replaced by His to perturb this hydrogen bond, showed that the ligand-free R to T transition rate was decreased by 20-fold compared with that for Hb A. This mutation caused the destabilization of the transition state in the R to T quaternary structure change by about 7 kJ mol-1, indicating that the hydrogen bond between Tyr-42 alpha and Asp-99 beta plays a definite role in the R-T transition as well as in stabilization of the equilibrium T state. Hb Phe-145 beta, in which Tyr-145 beta is replaced by Phe and the intrasubunit hydrogen bond between Tyr-145 beta and Val-98 beta is lacking, also showed a slow R-T transition rate as observed in Hb His-42 alpha. The published crystallographic data suggest that this intrasubunit hydrogen bond stabilizes the transition state by reducing the freedom of motion of the C-terminus of the beta subunit and, thereby, facilitates the R-T transition.

Amino Acid Sequence↗

PubMed1993

Structure, chromosomal localization, and expression of mouse reg genes, reg I and reg II. A novel type of reg gene, reg II, exists in the mouse genome.

We previously identified a gene, reg (i.e. regenerating gene), in the screening of a regenerating islet-derived cDNA library of rat (Terazono, K., Yamamoto, H., Takasawa, S., Shiga, K., Yonemura, Y., Tochino, Y., and Okamoto, H. (1988) J. Biol. Chem. 263, 2111-2114), and isolated a human reg cDNA and gene (Watanabe, T., Yonekura, H., Terazono, K., Yamamoto, H., and Okamoto, H. (1990) J. Biol. Chem. 265, 7432-7439); the rat and human cDNAs encode 165- and 166-amino acid proteins, respectively. Until now, it was thought that there is a single locus for Reg protein in the mammalian genome. In this study, we isolated two distinct cDNAs and genes, one of which was a mouse homologue to rat and human reg gene, the other a novel type of reg gene. We designated them reg I and reg II, respectively. The two proteins encoded by these genes share 76% amino acid sequence identity with each other. Both genes span about 3 kilobase pairs, and the genomic organization of six exons and five introns is conserved between them. Chromosomal mapping studies indicate that the reg I gene is localized on mouse chromosome 12, whereas the reg II gene is localized on chromosome 3. By Northern blot analysis, both reg I and reg II mRNAs are detected in the normal pancreas and hyperplastic islets of aurothioglucose-treated mice, but not in the normal islets. It is remarkable that in the gallbladder reg I is expressed, but reg II is not.

Amino Acid Sequence↗

PubMed1993

Obstructive jaundice associated with extrahepatic portal vein obstruction: report of two cases.

We herein report two cases of obstructive jaundice with markedly dilated collateral veins either in or around the bile duct in the setting of extrahepatic portal vein obstruction (EHPO). In the first case, a proximal splenorenal shunt provided relief of biliary stenosis as well as eradication of esophageal varices due to a decompression of portal hypertension. This evidence proved that the markedly extended collateral veins in the hepatoduodenal ligament caused biliary stenosis by compressing the bile duct. In the second case, obstructive jaundice was probably caused by cholangitis and was relieved with biliary drainage. Portal decompressive surgery was not indicated because of the slight degree of esophageal varices. The relationship between cholangitis and EHPO in these patients calls for further investigation. In cases with EHPO manifesting obstructive jaundice associated with risky esophageal varices, portal decompressive surgery is recommended as the procedure of choice.

Adolescent↗

PubMed1993

Ischemic heart disease showing unusual angiographic findings.

Atherosclerotic lesions usually occur in the proximal and middle portion of the coronary arteries. Multiple obstructive lesions appearing only in the peripheral branches without lesions in the proximal or distal portion have not been reported. We encountered a patient with ischemic heart disease showing multiple obstruction in the peripheral branches of the right and left coronary arteries without significant stenotic lesions in the proximal or middle portion. This 49-year-old male was admitted to Yamada Red Cross Hospital due to angina pectoris. Coronary risk factors for him included hypertension, abnormal glucose tolerance, smoking habit, and obesity. Laboratory studies showed a complete blood count and normal blood chemistries, as well as thromboplastin and prothrombin times. Coronary angiography showed multiple obstruction or marked stenosis in the distal portion and peripheral branches; there was no stenosis in the proximal and middle portions. Left ventriculography showed severe hypokinesis in the diaphragmatic segment. Biopsy of the left ventricular endocardium showed interstitial fibrosis but showed no abnormalities in the myocardial fibers or cell infiltration to perivascular areas and vascular walls. Coronary angiography after two months showed multiple lesions, as previously observed. Although ischemic heart disease is caused by various types of vasculitis, embolism, coronary spasm, and fibromuscular dysplasia, in this patient, there were no findings suggestive of causes other than atherosclerosis. This case is interesting in terms of rare angiographic findings and its cause.

Coronary Angiography↗

PubMed1993

A case of dilated cardiomyopathy manifested by exercise-induced left bundle branch block.

We report an extremely rare case of dilated cardiomyopathy manifested by exercise-induced left bundle branch block. A 63-year-old female came to our hospital because supraventricular arrhythmia had been detected at a check-up. A treadmill exercise test induced left bundle branch block. However, chest X-ray and echocardiography revealed no abnormal finding. Two years later, the patient experienced exertional dyspnea. A chest X-ray examination showed cardiomegaly, and echocardiography showed a moderate impairment of left ventricular function with left ventricular dilatation. The diagnosis of dilated cardiomyopathy was made by left ventricular myocardial biopsy. No previous cases have initially shown exercise-induced left bundle branch block which was followed by left ventricular dysfunction due to dilated cardiomyopathy. Careful long-term observation of the clinical course is necessary in patients with exercise-induced left bundle branch block which shows no significant underlying disease.

Bundle-Branch Block↗

PubMed1992

The primary structure of rat rig/ribosomal protein S15 gene.

We have isolated rat rig/ribosomal protein S15 gene from a DNA library derived from a rat insulinoma and determined the complete nucleotide sequence. The rat rig/S15 gene is composed of four exons and three introns spanning 2 kbp and exhibits distinctive structural features unique for a ribosomal protein gene.

Amino Acid Sequence↗

PubMed1992

Structural determination of Saccharomyces cerevisiae rig gene and identification of its product as ribosomal protein S21.

rig was originally isolated from a rat insulinoma-derived cDNA library. The 145 amino acid sequence of the rig protein is invariant in mammalian cDNAs. In this paper, we have isolated the cDNA and genomic clones for yeast (Saccharomyces cerevisiae) rig, determined their nucleotide sequences, and identified the gene product. The gene and the mRNA encode a basic protein of 142 amino acids which has 61.3% amino acid identity with mammalian rig protein. On two-dimensional gel electrophoresis, the in vitro transcription/translation product of yeast rig cDNA co-migrated with yeast ribosomal protein S21. These results led to the conclusion that yeast rig ribosomal protein S21 and to the determination of the previously unknown primary structure of yeast S21 protein. Unlike most ribosomal protein genes of S. cerevisiae, the gene exists as a single copy in a haploid set of the yeast genome and has no intron, locating at chromosome VII or XV.

Amino Acid Sequence↗

PubMed1992

Islet beta-cell regeneration and reg genes.

In this paper, we show that the reg gene is expressed in experimentally induced regenerating or hyperplastic islets. We have previously reported that ectopic expression of the reg gene occurs in some human colonic and rectal tumors, suggesting that enhanced reg expression may be related to the proliferative state of tumor cells. Reg protein has also been shown to have significant sequence homology with plant and animal lectins, a class of compounds that has been shown to be a growth promoter. At present, any direct relationship between reg protein and beta-cell replication remains to be established. However, since the reg protein is a secretory protein and reg can be expressed at an early stage of pancreatic cell differentiation, the reg protein may act on the stem cells of beta-cells in an autocrine or paracrine manner. In normal mature exocrine cells, the reg gene is expressed and the gene product may be necessary to maintain adequate exocrine pancreatic function. The physiological reasons for the maintenance of two functional reg genes encoding proteins of slightly different sequence in mice are still unclear. The cloning of additional rat and human non-allelic reg genes could provide additional clues.

Alloxan↗

PubMed1992

[A case of congenital long QT syndrome associated with T wave alternans].

A case was presented in which a rare T wave alternans occurred in association with congenital long QT syndrome. A 71-year-old woman, who had experienced several syncopal attacks per year for the previous forty years, was admitted for further evaluation of the syncope. She had a family history of sudden death (sister) and QT prolongation (son). Electrocardiogram showed a corrected QT interval of 0.68 seconds. Treadmill exercise-tolerance test revealed both T wave alternans immediately after exercise and torsades de pointes 150 seconds after exercise. The syncope was induced by the mental excitation. A prolonged corrected QT interval reduced from 0.70 seconds to 0.58 seconds by the correction of her serum potassium and magnesium. The effect of propranolol, verapamil, phenytoin or mexiletine on T wave alternans and ventricular arrhythmia was evaluated by the treadmill exercise-tolerance test. The treatment with propranolol was most effective.

Adrenergic beta-Antagonists↗

PubMed1992

[Exercise-induced left bundle branch block in treadmill exercise test: clinical significance and prognosis].

Exercise induced left bundle branch block (LBBB) is poorly understood. We investigated its clinical characteristics and prognosis. The records of 3,459 consecutive patients who underwent treadmill exercise testing were reviewed. Exercise induced LBBB was identified in 6 patients (0.17%). Three out of six patients had underlying cardiac disease; two had coronary artery disease and one dilated cardiomyopathy. Three out of five had redistribution on Tl-201 myocardial scintigraphy. In one of these three patients, however, coronary angiography demonstrated no significant narrowings. Tl-201 myocardial scintigraphy in patients with exercise induced LBBB may be undetermined for coronary artery disease. During follow-up, one of six patients had depressed left ventricular function and was diagnosed as having dilated cardiomyopathy. It is thus obvious that exercise induced LBBB without a specific underlying heart disease should be followed up carefully.

Aged↗

PubMed1991

Pressure effects on carbon monoxide rebinding to the isolated alpha and beta chains of human hemoglobin.

The effects of pressure on the recombination kinetics of carbon monoxide binding to the isolated alpha and beta chains of human adult hemoglobin at pH 7, approximately 20 degrees C, were studied by the use of millisecond and nanosecond laser photolyses. The kinetic data were analyzed on the basis of a simple three-species model, which assumes two elementary reaction processes of bond formation and ligand migration steps. The activation volume for each elementary step was obtained from the pressure dependence of the rate constants. A pressure-dependent activation volume change from negative to positive values in the bimolecular carbon monoxide association reaction was observed for both of the isolated chains. This finding is attributed to a change of the rate-limiting step from the bond formation step to the ligand migration step. For both of the isolated chains, the activation volumes for ligand migration into and from the protein were estimated as +12-16 and +7-11 cm3 mol-1, respectively. These positive activation volumes for the ligand migration process may be caused by conformational fluctuations of proteins, that is, the conformational changes from "closed" to "open" structure. In the iron-ligand bond formation process, the activation volumes are -15 to -22 cm3 mol-1, which are almost identical to that for the model heme complexes [Taube, D. J., Projahn, H.-D., van Eldik, R., Magde, D., & Traylor, T. G. (1990) J. Am. Chem. Soc. 112, 6880-6886]. Accordingly, the surrounding protein contributions to the activation volumes for the bond formation process could be small.(ABSTRACT TRUNCATED AT 250 WORDS)

Carbon Monoxide↗