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Biomedical subjects

Y Nishiya

Publications and source records attributed to Y Nishiya.

At least 37 records · Page 2Linked to original sources

Identification of an insertion sequence, IS1203 variant, in a Shiga toxin 2 gene of Escherichia coli O157:H7.

An insertion sequence composed of 1310 bp was found in Shiga toxin 2 genes of some isolates of Escherichia coli O157:H7. This insertion sequence showed extremely high homology with IS1203 of E. coli O111:H(-). This IS1203 variant was inserted in the region encoding the amino-terminus of the B subunit with a duplication of 3 bp at the target site, resulting in inactivation of the Shiga toxin 2 gene.

Journal Article↗

The guanosine binding mechanism of the Tetrahymena group I intron.

The Tetrahymena group I intron catalyzes self-splicing through two consecutive transesterification reactions, using a single guanosine-binding site (GBS). In this study, we constructed a model RNA that contains the GBS and a conserved guanosine nucleotide at the 3'-terminus of the intron (omegaG). We determined by NMR the solution structure of this model RNA, and revealed the guanosine binding mechanism of the group I intron. The G22 residue, corresponding to omegaG, participates in a base triple, G22 xx G3 x C12, hydrogen-bonding to the major groove edge of the Watson-Crick G3 x C12 pair. The G22 residue also interacts with A2, which is semi-conserved in all sequenced group I introns.

Animals↗

[Tricuspid valve replacement in a patient with idiopathic tricuspid regurgitation].

Tricuspid regurgitation is often seen but is rarely categorized as "idiopathic". A 66-year-old man suspected of idiopathic tricuspid regurgitation underwent tricuspid valve replacement. With right heart failure over twenty years, his CTR increased to eighty six (86) per cent. During operation, no abnormal findings were observed except for the dilated tricuspid valve ring. Then a stented porcine xenograft (Carpentier-Edwards 33 mm) was inserted in the tricuspid valve annulus. Seven years after replacement, he died. Autopsy demonstrated myocardial hypertrophy of the right ventricle and dilatation of the right heart, supporting the diagnosis of idiopathic tricuspid regurgitation.

Aged↗

Purification and characterization of a novel glycine oxidase from Bacillus subtilis.

The open reading frame yjbR which had been sequenced as a part of the Bacillus subtilis genome project encodes a putative 40.9-kDa protein. The yjbR-coding sequence was slightly similar to those of bacterial sarcosine oxidases and possibly compatible with the tertiary structure of the porcine kidney D-amino acid oxidase. The yjbR gene product was overproduced in Escherichia coli, purified to homogeneity from the recombinant strain, and characterized. This protein effectively catalyzed the oxidation of sarcosine (N-methylglycine), N-ethylglycine and glycine. Lower activities on D-alanine, D-valine, and D-proline were detected although no activities were shown on L-amino acids and other D-amino acids. Since glycine is a product and not a substrate for sarcosine oxidase, this protein is not a type of demethylating enzymes but a novel deaminating oxidase, named glycine oxidase as a common name. Several enzymatic properties of the B. subtilis glycine oxidase were also investigated.

Amino Acid Oxidoreductases↗

Thermostable glycerol kinase from Thermus flavus: cloning, sequencing, and expression of the enzyme gene.

The thermostable glycerol kinase (EC 2.7.1.30) gene from Thermus flavus was cloned and expressed in Escherichia coli DH5 alpha. An open reading frame of 1488 bp for the glycerol kinase gene (glpK) starting with an ATG methionine codon was found, which encodes a protein of 496 amino acid residues whose calculated molecular weight is 54,835. The amino acid sequence of T. flavus glycerol kinase is 80.6% and 64.1% identical with those of Bacillus subtilis and E. coli. Transformants of E. coli DH5 alpha harboring plasmid pGYK12 with a 1505 bp chromosomal DNA fragment containing the T. flavus glycerol kinase gene showed about 23.8-fold higher glycerol kinase activity than T. flavus.

Amino Acid Sequence↗

A new method for right ventricular endomyocardial biopsy via the femoral veins: a novel approach employing a right ventriculography catheter (Nishiya type).

We have devised a new method for right ventricular endomyocardial biopsy, in which the use of a right ventriculography catheter (Nishiya Type) allows us to ensure the introduction of a guiding sheath (Cordis, right-angled long sheath) to the apical portion of the right ventricle by way of the femoral vein. With this method, we have biopsied the endomyocardium in 52 patients during the last 4 years. In each case, it only took us several minutes to complete the biopsy procedure. Neither failed applications nor significant complications were seen. We believe that this technique provides a clinically beneficial means as it enables us to readily and safely achieve right ventricular endomyocardial biopsy.

Adolescent↗

Gene cluster for creatinine degradation in Arthrobacter sp. TE1826.

The genes encoding creatininase (CrnA; 258 residues) and creatinase (CreA; 411 residues) from Arthrobacter sp. TE1826 were cloned and sequenced. The genes form a cluster with the sarcosine oxidase gene (soxA) and its regulator gene (soxR), which were cloned previously. The deduced amino acid sequences of CrnA and CreA show 35.9% and 63.1% identity, respectively to the corresponding Pseudomonas enzymes. CrnA and CreA were purified from the recombinant strains and characterized. Other open reading frames (creB and crnB), encoding proteins similar to several transporters, were found downstream of creA and crnA, respectively.

Amidohydrolases↗

Effect of non-ionic detergents on apparent enzyme mechanism: V121A mutant of Streptomyces cholesterol oxidase endowed with enhanced sensitivity towards detergents.

One of the mutants of Streptomyces cholesterol oxidase with the Val121Ala mutation (V121A) was kinetically analysed. Although the reaction rate-substrate concentration curve of wild type follows a simple Michaelis-Menten equation, that of V121A is sigmoidal. The cooperativity was apparent and caused by non-ionic detergents that were used as a solvent of cholesterol. The concentration dependence of V121A on detergents was more significant than that of wild type, although the reaction rates of both enzymes decrease as the concentrations of detergents increase. Further experiments suggested that less hydrophobic interactions between V121A and detergents should be responsible for the apparent cooperativity. Since Val121 is in a hydrophobic loop located near the active site, the mutational effect is structurally discussed.

Binding Sites↗

A new preshaped catheter for right ventriculography with minimal artificial tricuspid regurgitation.

A new type of preshaped angiographic catheter for right ventriculography to be inserted from the femoral vein has been developed. Right ventriculography using this catheter was conducted in about five hundred clinical cases. The result has revealed that this catheter is very useful in clinical settings without causing catheter-induced tricuspid regurgitation (TR) allowing easier and safer right ventriculography as compared with conventional catheters.

Cardiac Catheterization↗

[A case of mediastinal teratoma discovered availing of pneumonia].

A 42-year-old woman was admitted to our hospital with hemosputum and an abnormal shadow on chest X-ray. Although her hemosputum disappeared after the treatment of pneumonia, the abnormal shadow was not improved. Chest CT showed a mediastinal mass. She underwent thoracotomy and the tumor, measuring 6 x 5 x 5 cm in size, was resected. Pathological diagnosis was a mature type teratoma with direct invasion to the lung. We have to take mediastinal teratoma into consideration as one of the causes of pneumonia.

Adult↗

Enzymatic assay for chloride ion with chloride-dependent sarcosine oxidase created by site-directed mutagenesis.

Serum chloride ion is routinely assayed in clinical laboratories. We have developed a new enzymatic assay for determining the chloride ion concentration. The method involves the use of a mutant sarcosine oxidase, which was created as desired by site-directed mutagenesis and showed chloride-dependent activity. The enzyme which is reactivated by the chloride ion forms hydrogen peroxide from sarcosine. The production of hydrogen peroxide is measured by the 4-amino-antipyrine peroxidase system. The increase of the reaction rate was proportional to the chloride ion concentration. A lag time of the time course was not observed, and the reaction rate for a blank was not detected. Therefore, a rate assay could be adopted. A standard curve of the assay reagent was linear up to 180 mM chloride ion of the sample. The specificity for the bromide ion was 43% of that of the chloride ion, although it was 0% for other ion species. When serum samples were used, within-day coefficient variations (CVs) and day-to-day CVs were below 1.5%. A good correlation with the comparison assay was observed by using 160 samples of normal and abnormal patient sera. This method can easily and reliably be used for the accurate determination of chloride ion concentration in serum or other samples.

Amino Acid Sequence↗

Improvement of thermal stability of Streptomyces cholesterol oxidase by random mutagenesis and a structural interpretation.

Random mutagenesis was used to enhance the thermal stability of Streptomyces cholesterol oxidase. Four thermostable mutants were isolated and the following amino acid substitutions were identified: Ser103 to Thr (mutant S103T), Val121 to Ala (mutant V121A), Arg135 to His (mutant R135H) and Val145 to Glu (mutant V145E). The wild-type and mutant enzymes were purified and characterized. The properties of mutants S103T, V121A and R135H were similar to those of the wild type but they showed improved thermal stability. When the V145E mutation was introduced, the thermal stability of the enzyme was markedly increased and the optimum pH was desirably changed to encompass a broad range from acid to alkali. Analysis of multiple mutants constructed by site-directed mutagenesis showed that all the mutations except that of R135H had an additive influence on the other mutations. These mutational effects are discussed in terms of a three-dimensional structural model of the enzyme constructed on the basis of homology modelling.

Cholesterol↗

[A case of anastomotic aneurysm-enteric fistula].

We successfully performed circulation reconstructive surgery and repair of a small intestine resection, primarily at an anatomical position, for perforation of a pseudoaneurysm in the small intestine, which occurred in an anastomosis site of the right leg of a Y graft of an artificial blood vessel and internal iliac artery graft on a patient, at the remote time of 18 years after she had received a Y-type artificial blood vessel. The patient's life was saved. The results suggest that it is important to make a definitive diagnosis quickly after the occurrence of symptoms and to select an appropriate surgical technique according to the findings of infection adjacent to the perforation site.

Aged↗

Analysis of interaction between the Arthrobacter sarcosine oxidase and the coenzyme flavin adenine dinucleotide by site-directed mutagenesis.

Sarcosine oxidase from Arthrobacter sp. TE1826 (SoxA) tightly binds with the coenzyme flavin adenine dinucleotide (FAD). The amino-terminal region of this enzyme was recognized as a part of the FAD-binding domain by homology search analysis. Comparison with other structurally well-known flavoproteins suggested that the aspartate residue at position 35 (D-35) and the motif sequence (six residues at positions 12 to 17) were important for the interaction with FAD. Site-directed mutagenesis of each position was performed, and mutant SoxAs were purified and characterized. When D-35 was substituted with glutamate, asparagine, and alanine, it was indicated that the carboxyl group of the side chain interacted with FAD. Changes in the enzyme-bound FAD were also observed from the altered spectral profiles. Thirteen mutant SoxAs were obtained by replacing amino acids in the motif sequence. Most of them showed inhibited or remarkably decreased sarcosine oxidase activity, and their spectral profiles were altered. However, some of them were reactivated by chloride ion. Their spectral profiles also became close to that of wild type in the presence of chloride ion. These results strongly suggest that the inhibition of interaction of enzyme with FAD was caused by the substitution in the motif and that it could be recovered under different conditions.

Amino Acid Sequence↗

Active site analysis and stabilization of sarcosine oxidase by the substitution of cysteine residues.

Two cysteine residues (C-265 and C-318) in the putative hydrophilic regions of sarcosine oxidase were substituted by using site-directed mutagenesis. Since the mutant with the C-to-S mutation at position 318 (C318S) lost the enzyme activity, C-318 (conserved among sarcosine oxidases) is most likely a part of the active site. C265S, C265A, C265D, and C265R showed nearly the same enzymatic properties as those of the wild type. However, they were much more stable than the wild type in the presence of inhibitors that modified the thiol group. Moreover, they were extremely stable throughout the cultivation of the recombinant strains or even in cell extracts.

Arthrobacter↗

[Cavernostomy and muscle plombage with the pedicled latissimus dorsi and serratus anterior flap for recurrent large bullae with infection: a case report].

A 70-year-old male, who suffered from infection by MRSA and Aspergillus of the recurrent large bullae, was operated on cavernostomy for space sterilization. About three months later he underwent muscle plombage with the pedicled latissimus dorsi and serratus anterior flap. These muscles are ideal to place in open wound in the thorax because of the following reasons: These have the ability to eliminate local infection, a sufficient length and volume to obliterate an open cavity. And transposition of these muscles cause minimal functional upper extremity disability. Therefore, we considered that this technic was useful for the case of an infectious intrathoracic cavity with many bronchial fistulas.

Aged↗

Primary structure of the ADE1 gene from Candida utilis.

The ADE1 gene from Candida utilis CA(u)-37, a strain used for commercially producing enzymes, was cloned by complementation of the ade1 mutation of Saccharomyces cerevisiae. It was composed of 903 bp, and the deduced amino acid sequence was 70% homologous to those of the ADE1 genes of S. cerevisiae and Candida maltosa. The highly preserved region of SAICAR synthetase, the ADE1 gene product, was also found by a homology search.

Amino Acid Sequence↗