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Biomedical subjects

I Matsui

Publications and source records attributed to I Matsui.

At least 19 recordsLinked to original sources
PubMed1992

A mutant alpha-amylase with enhanced activity specific for short substrates.

The 210th lysine (K210) at the active site in Saccharomycopsis fibuligera alpha-amylase was altered to arginine (R) or asparagine (N) by site-directed mutagenesis. Replacement of K210 by R strengthened the 7th and weakened the 8th subsite affinities. K210 was found to contribute to both the 8th and the 7th subsites. The catalytic activity of the K210R enzyme for the hydrolysis of maltose (G2) was three-times higher than that of the native enzyme due to an increase in the affinity of the 7th subsite adjacent to the catalytic site, whereas the activity of the K210N enzyme for G2 was decreased to 1% of that of the native enzyme by a reduction in the 7th subsite affinity.

Amylose↗
PubMed1992

Alteration of bond-cleavage pattern in the hydrolysis catalyzed by Saccharomycopsis alpha-amylase altered by site-directed mutagenesis.

The 210th lysine (K) residue in the Saccharomycopsis alpha-amylase (Sfamy) molecule was replaced by arginine (R) and asparagine (N) residues by site-directed mutagenesis. The influences of the replacements on the bond-cleavage pattern for several substrates were analyzed. Both mutant enzymes, K210R and K210N, cleave mainly the first glycosidic bond from the reducing end of maltotetraose (G4), while the native enzyme hydrolyzes mainly the second bond from the reducing end. We changed successfully the major cleavage point in the hydrolysis reaction of G4. The 8th subsite affinities of the K210R and K210N enzymes are calculated to be +2.52 and -0.01 kcal/mol, respectively, whereas that of the native enzyme is +3.32 kcal/mol as reported in the previous paper. These affinity values suggest that the K210 residue composes the 8th subsite, one of major subsites, and that a positively charged amino residue is necessary for the 8th subsite affinity. The K210N enzyme is found to be less active for short substrates like maltotetraose (G4) than for long substrates like amylose A (approximately G18). The reduced catalytic activity specifically for the short substrates is also attributable to the remarkable decrease in the affinity of the 8th subsite.

Amino Acid Sequence↗
PubMed1992

Multi-functional roles of a histidine residue in human pancreatic alpha-amylase.

Functional roles of histidine residues at the active site in human pancreatic alpha-amylase were examined by protein engineering. Three histidine residues at 101, 201, and 299 were converted to asparagine residues, respectively. It was found that His201 played multi-functional roles concerning so many functions; substrate binding, control of optimum pH, change in substrate specificity, activation by chloride ion, and inhibition by a proteinaceous inhibitor.

Binding Sites↗
PubMed1992

Biological activities of rabbit antibodies against synthetic human thyrotropin receptor peptides representing thyrotropin binding regions.

Recently, we have shown that the thyrotropin (TSH) binding regions of human thyrotropin receptor (TSHR) reside in two areas within residues 12-44 and 308-344. Serial antisera were raised against four overlapping synthetic peptides representing these two regions of TSHR (peptides 12-30, 24-44, 308-328, and 324-344) and were investigated for their ability to stimulate or block the cultured porcine thyroid cells. In addition, serum concentrations of triiodothyronine (T3) and thyroxine (T4) in serial sera obtained from each rabbit were examined. It was shown that residues of 12-30 and 324-344 of TSHR, respectively, are the site (at least a part of the site) where stimulating (TSAb) and blocking type (TSBAb) immunoglobulins are directed.

Amino Acid Sequence↗
PubMed1992

Non-suppressed thyroidal radioactive iodine uptake (RAIU) in thyrotoxic phase in a case of subacute thyroiditis with thyroid-stimulating antibodies (TSAb).

In this paper, we report a 49-year-old female with subacute thyroiditis who had thyroid-stimulating antibodies (TSAb) and thyroid-stimulation-blocking antibodies (TSBAb) in serum. Although she was in the thyrotoxic phase and TSH was suppressed in May, 1990, her radioactive iodine uptake (RAIU) was not suppressed (35.5%) and a thyroid scan disclosed a diffuse goiter with no defect. Serum assays revealed the presence of TSAb, but TSBAb were negative. In August, 1990, the right lobe became undetectable by thyroid scan when the RAIU was 20.7% with the TSH level remaining suppressed. At that time, TSAb were negative, while TSBAb were positive. When the RAIU was 31.1% in October, 1990, both thyroid lobes became visible and the TSH level was normalized. TSBAb became negative, and although TSAb reappeared it later became undetectable. These results indicate that the changes in the patient's thyroid scan and RAIU were attributable to the presence of TSAb.

Autoantibodies↗
PubMed1991

The pH dependence of the action pattern in porcine pancreatic alpha-amylase-catalyzed reaction for maltooligosaccharide substrates.

Porcine pancreatic alpha-amylase (EC 3.2.1.1; abbreviated PPA), which hydrolyzes alpha-D-(1,4) glucosidic bonds in starch and amylose, displays an optimum at pH 6.9 for the majority of substrates. The optimum pH, however, shifted to 5.2 for the hydrolysis of some low molecular substrates (Ishikawa, K., et al., 1990, Biochemistry 29, 7119-7123). Details of the substrate-dependent shift of the optimum pH in PPA were studied by use of a series of maltooligosaccharides with 14C-labeled reducing end glucose as substrates. The optimum pH for maltotriose was 5.2, whereas that for maltopentaose and maltohexaose was unchanged at pH 6.9. The pH profile for the intermediate size substrate maltotetraose showed abnormality; the apparent optimum pH was broadened between 5.5 and 6.5 and the bond cleavage pattern depended on pH, unlike that for the other substrates examined. These results were independent of either buffer systems or substrate concentration. Analyses of the hydrolysates of the maltooligosaccharides revealed that the shift of the optimum pH to the neutral region occurred only when the fifth subsite of PPA in the productive binding modes was occupied by a glucosyl residue of a substrate. The three-catalytic residue model of PPA deduced from the analysis of the hydrolysis of some modified maltooligosaccharides (p-nitrophenyl-alpha-D-maltoside, gamma-cyclodextrin, maltopentaitol, and maltohexaitol) (Ishikawa, K., et al., 1990, Biochemistry 29, 7119-7123) was successfully adapted to the linear maltooligosaccharides used in this work. These results indicate that the different productive binding modes of the linear oligosaccharide substrates affect directly the catalytic power and the optimum pH of PPA.

Animals↗
PubMed1991

An increase in the transglycosylation activity of Saccharomycopsis alpha-amylase altered by site-directed mutagenesis.

The 84th tryptophan residue in Saccharomycopsis alpha-amylase molecule was replaced by a leucine residue and the resulting site-directed mutant, W84L enzyme, showed an increase in transglycosylation activity. At a 40% digestion point of maltoheptaose (G7), for example, maltooligosaccharide products larger than maltodecaose (G10) amounted to approx. 60% of the total product from the mutant enzyme reaction, whereas no such large products were observed in the native enzyme reaction. Analysis of the reaction products from p-nitrophenyl maltooligosaccharides indicated that these large products were formed by addition of the hydrolysis products on the nonreducing end side to the starting intact substrates. These results suggest that the tryptophan residue located at subsite 3 of the enzyme plays an important role not only to hold the substrate, but also to liberate the hydrolysis products from the substrate binding pocket.

Glycosylation↗
PubMed1991

Alpha 1-adrenergic regulation of thyrotropin-stimulated release of 3, 5, 3' -triiodothyronine and thyroxine from perifused mouse thyroid.

The effect of methoxamine, a specific alpha 1-adrenergic agonist, on the release of T3, T4 and cAMP from perifused mouse thyroid was studied to clarify the role of the alpha 1-adrenergic receptor in the regulation of thyroid hormone secretion. TSH-stimulated T3 and T4 release was inhibited significantly by methoxamine. With regard to cAMP release, methoxamine inhibited TSH-stimulated cAMP release in the presence of 4-(3-butoxy-4-methoxybenzyl)-2-imidazolidinone but did not inhibit TSH-stimulated cAMP release in the presence of 3-isobutyl-1-methylxanthine. Methoxamine did significantly suppress TSH-stimulated release of T3 and T4 in the presence of each phosphodiesterase inhibitor. Depletion of Ca2+ in the perifusion buffer abolished completely the inhibitory effect of methoxamine on TSH-stimulated T3 and T4 release. The present study suggests that activation of the alpha 1-adrenergic receptor inhibits TSH-stimulated T3 and T4 secretion through a Ca(2+)-dependent mechanism in the mouse thyroid gland.

1-Methyl-3-isobutylxanthine↗
PubMed1991

Subsite structure of Saccharomycopsis alpha-amylase secreted from Saccharomyces cerevisiae.

The kinetic parameters (kcat/Km) and the cleaved-bond distributions for the hydrolysis of linear maltooligosaccharides Gn (3 less than or equal to n less than or equal to 9) by Saccharomycopsis alpha-amylase (Sfamy) secreted from Saccharomyces cerevisiae were determined at pH 5.25 and 25 degrees C. The subsite affinities of Sfamy were also evaluated from these data. The subsite structure of Sfamy is characteristic of the active site of an endo-cleavage type enzyme, consisting of internal repulsive sites with the catalytic residues and external attractive sites. Moreover, the pKa values of the catalytic residues were calculated from the pH dependence plot of the kinetic parameter (kcat/Km). The amino acid residues which contribute to the subsite affinities and the catalytic activity of Sfamy are proposed and compared with those of Taka-amylase A.

Kinetics↗
PubMed1991

Inhibitory effects of ionophore A23187 on the release of thyroid hormone and colloid reabsorption in mouse thyroid glands.

The effect of the ionophore A23187 on a. the release of thyroid hormone from perifused mouse thyroid glands and b. the morphological changes in follicular epithelial cells was evaluated. A23187 at a concentration of 5 mumol/l significantly inhibited both the TSH- and the forskolin-stimulated release of T3 and T4. In the presence of 3-isobutyl-1-methylxanthine or (4-(3-butoxy-4-methoxybenzyl)-2-imidazolidinone) RO 20-1724, A23187 did not affect the forskolin-stimulated release of cAMP, but did inhibit the release of T3 and T4 stimulated by forskolin. Light and electron microscopic evaluation of the follicular epithelial cells of mouse thyroid tissues following 1-h stimulation with forskolin showed numerous pseudopods engulfing luminal colloid of various size and the presence of reabsorbed colloid droplets in the apical cytoplasm. Quantitative electron microscopic analysis revealed that the addition of the ionophore A23187 reduced the number of reabsorbed colloid droplets to one eighth in follicular epithelial cells. These observations suggest that the increase in intracellular Ca2+ induced by the ionophore A23187 inhibits the TSH-stimulated thyroid hormone release independently of the cAMP level, and that the suppression of thyroid hormone release may be due to an inhibition of colloid reabsorption.

1-Methyl-3-isobutylxanthine↗
PubMed1990

Child abuse of one of a pair of twins in Japan.

A nationwide survey in Japan on child abuse and neglect revealed that 10% of the victims were products of multiple births. None of the victims who were singletons had multiple-birth siblings, and only in a few cases were both twins abused. The findings indicated that one rather than both of a pair of twins was likely to be abused in Japan. Abuse of both twins was likely when there were serious parental or family problems, whereas abuse of one twin was associated with the child's medical problems or non-home care. There was no instance of abuse of a pair of twins when both were handicapped. Comparisons of the abused twin with the non-abused co-twin and examination of the abuser's attitude to the victim suggested that the difference between twins in their development or in their response to parents increased the stress of child-rearing and encouraged favouritism, which resulted in abuse of only one twin. Comparison by parents of children with their siblings may be a common factor in general child abuse because it is a natural thing for parents to do.

Adult↗
PubMed1990

Substrate-dependent shift of optimum pH in porcine pancreatic alpha-amylase-catalyzed reactions.

Porcine pancreatic alpha-amylase (EC 3.2.1.1, abbreviated as PPA) hydrolyzes alpha-D-(1,4) glucosidic bonds in starch and amylose at random, and the optimum pH for the substrates is 6.9. The optimum pH, however, shifted to 5.2 for the hydrolytic reaction of low molecular weight oligosaccharide substrates such as p-nitrophenyl alpha-D-maltoside, gamma-cyclodextrin, maltotetaitol, and maltopentaitol. The optimum pH for the oligosaccharides consisting of more than five glucose residues, such as maltopentaose and maltohexaitol, was 6.9. From the analysis of the hydrolysates, it was clear that the shift of the optimum pH occurred only when the fifth subsite of PPA in the productive binding modes was occupied by a glucosyl residue of the substrates. The value of Km was independent of pH between 4 and 10 but that of kcat was dependent on pH. The pH profiles of kcat for the above substrates did not fit a simple bell-shaped curve predicted by a two-catalytic-group mechanism. Instead, they were well analyzed theoretically by three pK values and two intrinsic kcat values. Enthalpy changes for the three pK's (4.90, 5.35, and 8.55 at 30 degrees C) were determined from the temperature dependence of pH profiles for maltopentaitol and maltohexaitol to be 0.0, 2.87, and 7.33 kcal/mol, respectively. These results indicate that productive binding modes of the substrates directly affect the catalytic function of the enzyme. From the present thermodynamic analysis and reported three dimensional structure at the active site of PPA [Buisson, G. (1987) EMBO J. 6, 3909-3916], one can assume that a histidyl residue (101, 201, or 299) acts as a proton donor and two carboxyl groups (Asp 197, Glu 233, or Asp 300) act as proton donors or acceptors, and the productive binding mode covering the fifth subsite changes configurations between the catalytic residues and the glucosidic bond hydrolyzed and modulates kinetic parameters depending on pH.

Amylose↗
PubMed1990

The enzymatic and molecular characteristics of Saccharomycopsis alpha-amylase secreted from Saccharomyces cerevisiae.

The Saccharomycopsis fibuligera alpha-amylase (Sfamy) gene was expressed in Saccharomyces cerevisiae. The highest productivity of Sfamy was 70 mg per liter of culture broth. We purified Sfamy from the culture broth and identified the NH2 terminal primary sequence. This sequence suggests that the Sfamy gene product is synthesized as a pre-pro-precursor, and the pro-sequence is cleaved after a Lys-Arg sequence with the calpain-like endopeptidase encode by the KEX2 gene, resulting in mature Sfamy protein composed of 468 amino acids. Furthermore, the enzyme Sfamy is a glycoprotein in which one N-linked sugar chain containing mannose residues is attached to the Asn residue at the 198 position. The Km and kcat values were 1.1 x 10(-4) M and 1.4 x 10(2) sec-1, respectively, using amylose (the degree of polymerization n = 18) as a substrate. Moreover, the secondary structure, the location of the secondary elements including alpha-helix, beta-sheet, and loop, and tertiary structure were predicted theoretically on the basis of the molecular structure of Aspergillus oryzae alpha-amylase. Taka-amylase A (TAA). These results indicate that Sfamy protein is composed of main (M) and C-terminal (C) domains. The molecular structure of M domain closely resembles that of TAA, but the C domain appears to be more compact than that of TAA because of deletions at three regions forming turns and one region forming alpha-helix.

Amino Acid Sequence↗
PubMed1990

Mass screening for neuroblastoma in Japan. Committee of the Neuroblastoma Mass Screening Study of the Japanese Society of Pediatric Oncology.

The present status of NB mass screening in Japan, which is the first national trial in the world, is presented. This program was conducted in cooperation with infants' mothers, local health centers, screening centers and hospitals. Three hundred and thirty-seven cases detected by the program, from the start in Kyoto in 1974 to the end of 1989, are analyzed. As many cases were detected at early stages, 97% of them were expected to be cured. Several clinical, technical and programmatic problems are also discussed.

Child↗
PubMed1990

[Present status of neuroblastoma mass screening in Japan. Neuroblastoma-Committee of the Japanese Childhood Cancer Society].

In 1973, mass screening program for 6-month old infants for early detection of neuroblastoma using a VMA spot test of a urine sample was initiated in Kyoto. In 1985, nation wide mass screening was initiated throughout the entire country and the Government has given the financial support to each district. In 1988, the Government recommended the institution of mass screening by quantitative measurements of VMA, HVA and creatinine using HPLC (high performance liquid chromatography), instead of the qualitative test of VMA alone. From 1974, at the time of initiation of mass screening for neuroblastoma to the end of October, 1989, 383 cases with this tumor have been discovered throughout the screening program. Three hundreds eighty three cases (88%) of them had been registered to the Neuroblastoma Committee of the Japanese Society of Pediatric Oncology. In this paper, the mass screening program was introduced and the 337 cases with this tumor detected by 6-month old screening were analyzed their clinical symptoms, findings, urinary VMA and HVA levels, primary sites, weights of primary tumor, histology, stages at diagnosis, metastatic sites, and the results of the treatment. Three hundreds twenty eight cases (97%) of them are expected to be cured. And we discussed clinical problems related to mass screening program for neuroblastoma, such as an increase of the incidence of infantile neuroblastomas detected by this program and the spontaneous regression.

Adolescent↗
PubMed1990

[A study of environmental factors before and during pregnancy in childhood malignancy].

To determine risk factors in childhood malignancy, we compared parental and fetal exposure rates of some environmental factors among major children's malignancies based on 2722 cases of the Japan Children's Cancer Registry Database. The rates of parental exposure to irradiation, chemicals, and maternal dosage before and during pregnancies and of maternal smoking and drinking habit during pregnancies were slightly but statistically significantly high in some blastomas in which loss of heterozygosity of the genes has been reported. On the other hand, there were no significant correlations with these factors in leukemia and malignant lymphoma, which have been reported to be related to special chromosomal translocations. These possible risk factors should be examined by experimental and more detailed epidemiological studies.

Alcohol Drinking↗