Biomedical subjects

T Horii

Publications and source records attributed to T Horii.

At least 109 records · Page 6Linked to original sources

PubMed1995

Interaction of Tyr103 and Tyr264 of the RecA protein with DNA and nucleotide cofactors. Fluorescence study of engineered proteins.

To obtain structural insight on the interaction of the RecA protein with nucleotide cofactors (ATP and ADP) and DNA, we have made two engineered RecA proteins, in which either Tyr103 or Tyr264 was replaced with tryptophan. The fluorescence of tryptophan residues (two/subunit) of wild-type RecA is not significantly altered upon the binding of cofactor or DNA. Therefore, any detectable fluorescence change of the engineered proteins could be directly related to interaction with the particular inserted tryptophan residue. The fluorescence of Trp103 is almost completely quenched upon ADP binding, supporting a stacking interaction of adenine base of ADP with Tyr103. By contrast, with ATP the quenching of fluorescence of Trp103 is not complete (75%), possibly indicating that there is no stacking interaction with ATP. Such a difference could explain the antagonistic effects of ATP and ADP. Both nucleotides partially quench the fluorescence of Trp264 (about 70%), confirming that this residue is in the vicinity of the cofactor-binding site. The binding of ssDNA also decreases the fluorescence of both Trp103 and Trp264, the degree of quenching depending upon base composition and decreasing in the following order: poly(dT) > poly(dI) > M13 ssDNA > poly(dA). This order coincides with that of the binding affinities of these polynucleotides to RecA reported by Cazenave et al. [Cazenave, C., Chabbert, M., Toulmé, J. J. & Hélène, C. (1984) Biochim. Biophys. Acta 781, 7-13]. This correlation supports the finding that a region very close to Tyr103 interacts with DNA.

Adenosine Diphosphate↗

PubMed1995

The DNA-binding site of the RecA protein. Photochemical cross-linking of Tyr103 to single-stranded DNA.

To investigate the DNA-binding site in the Escherichia coli RecA protein, RecA was covalently cross-linked to oligodeoxythymidine [p(dT)14] by irradiation with ultraviolet light. We identified the site of cross-linking of the protein when the RecA.p(dT)14 complex was formed in the absence of nucleotide cofactor as well as in the presence of adenosine 5'-[gamma-thio]triphosphate. When RecA.p(dT)14 complex formed without nucleotide cofactor was irradiated with ultraviolet light, a cross-linked peptide was found after digestion with Achromobactor lyticus protease I. Amino acid composition of the peptide was determined. The results indicated that the site of cross-linking was in the region spanning amino acid residues 89-106. Further digestion of the cross-linked fragment with Staphylococcus aureus V8 protease indicated that Tyr103 was the site of cross-linking. When the complex formed with adenosine 5'-[gamma-thio]triphosphate was irradiated with ultraviolet light, two cross-linked sites were detected, which were in the region of residues 89-106 and residues 178-183. These regions are far from the two disordered loops in the crystal structure, which were suggested to be DNA-binding sites by Story et al. [Story, R. M., Weber, T. W. & Steitz, T. A. (1992) Nature 355, 318-325].

Adenosine Triphosphate↗

PubMed1995

Phylogenetic relationships within Taenia taeniaeformis variants and other taeniid cestodes inferred from the nucleotide sequence of the cytochrome c oxidase subunit I gene.

Nucleotide sequence variations in a region of the mitochondrial cytochrome c oxidase subunit I (COI) gene (391 bp) were examined within seven species of the genus Taenia and two species of the genus Echinococcus, including ten isolates of T. taeniaeformis and six isolates of E. multilocularis. More than a 12% rate of nucleotide differences between taeniid species was found, allowing the species to be distinguished. In E. multilocularis, no sequence variation was observed among isolates, regardless of the host (gray red-backed vole, tundra vole, pig, Norway rat) or area (Japan, Alaska) from which each metacestode had been isolated. In contrast, six distinct sequences were detected among the ten T. taeniaeformis isolates examined. The level of nucleotide variation in the COI gene within T. taeniaeformis isolates except for one isolate from the gray red-backed vole (TtACR), which has been proposed as a distinct strain or a different species, was about 0.3%-4.1%, whereas the COI gene sequence for TtACR differed from those of the other isolates, with levels being 9.0%-9.5%. Phylogenetic trees were then inferred from these sequence data using two different algorithms.

Amino Acid Sequence↗

PubMed1995

Coronary artery reoperation through the left thoracotomy with hypothermic circulatory arrest.

BACKGROUND: The left thoracotomy approach to avoid injury of the patent old graft and the myocardium with mid sternal reentry at coronary artery reoperation. METHODS: The left thoracotomy approach was used in 13 patients. There were 11 men and 2 women with a mean age of 63 years, ranging from 39 to 75 years. Three patients were having their third coronary bypass operation. In 11 patients, distal anastomoses were performed under circulatory arrest with moderate hypothermia. In the other 2 patients, distal anastomoses were performed on a beating heart. No aortic cross-clamp was applied in all patients. The mean number of distal anastomoses was 1.8; the grafted vessels were 11 anterior descending, 3 diagonal, 8 circumflex, and 1 posterolateral coronary arteries. Used grafts were 17 saphenous veins, 4 left internal thoracic arteries, and 2 gastroepiploic arteries. Inflow sites of the free graft were descending aorta in 10 patients and left subclavian artery in 3 patients. RESULTS: All patients were alive and well at the mean follow-up of 16 months, and all grafts were patent. CONCLUSIONS: The left thoracotomy approach is safe and effective for reoperation on the left coronary artery system, and circulatory arrest is convenient and safe for performing distal anastomosis.

Adult↗

PubMed1995

Analysis of the DNA binding site of Escherichia coli RecA protein.

To investigate the DNA binding site of RecA protein, we constructed 15 recA mutants having alterations in the regions homologous to the other ssDNA binding proteins. The in vivo analyses showed that the mutational change at Arg243, Lys248, Tyr264, or simultaneously at Lys6 and Lys19, or Lys6 and Lys23 caused severe defects in the recA functions, while other mutational changes did not. Purified RecA-K6A-K23A (Lys6 and Lys23 changed to Ala and Ala, respectively) protein was indistinguishable from the wild-type RecA protein in its binding to DNA. However, the RecA-R243A (Arg243 changed to Ala) and RecA-Y264A (Tyr264 changed to Ala) proteins were defective in binding to both ss- and ds-DNA. In self-oligomerization property, RecA-R243A was proficient but RecA-Y264A was deficient, suggesting that the RecA-R243A protein had a defect in DNA binding site and the RecA-Y264A protein was defective in its interaction with the adjacent RecA molecule. The region of residues 243-257 including the Arg243 is highly homologous to the DNA binding motif in the ssDNA binding proteins, while the eukaryotic RecA homologues have a similar structure at the amino-terminal side proximal to the nucleotide binding core. The region of residues 243-257 would be a part of the DNA binding site. The other parts of this site would be the Tyr103 and the region of residues 178-183, which were cross-linked to ssDNA. These three regions lie in a line in the crystal structure.

Amino Acid Sequence↗

PubMed1995

Em18 and Em16, new serologic marker epitopes for alveolar echinococcosis in western blot analysis, are the only two epitopes recognized by commercially available weak positive (cut off) sera for Em2plus-ELISA.

The assay system for antibody responses against Em2, the most specific antigen for serodiagnosis of alveolar echinococcosis (AE), has been established by enzyme-linked immunosorbent assay (ELISA) but not by Western blot assay, since Em2 antigen is not protein but carbohydrate in nature. Recently we reported that previously undescribed protein epitopes, designated Em18 and Em16 due to their molecular weights, were good serologic markers for AE by Western blot analysis. It has been shown that Em18 and Em16 are the only two epitopes recognized by commercially available weak positive (cut off) sera for the Em2plus-ELISA.

Antibodies, Helminth↗

PubMed1995

Effect of pH on activities of novel beta-lactamases and beta-lactamase inhibitors against these beta-lactamases.

The effects of acidic conditions on activities of seven beta-lactamases--TEM-1 (class A), KOXY (class A), IMP-1 (class B), AmpC (class C), MOX-1 (class C), OXA-5 (class D), and PSE-2 (class D)--and their inhibitors were measured. The enzymatic activities of KOXY, IMP-1, and MOX-1 at pH 5.8 were slightly lower than those at pH 7.5. However, the activities of PSE-2 and OXA-5 were greatly reduced at pH 5.8. All of the beta-lactamase inhibitors tested had poorer inhibitory activities at pH 5.8 than at pH 7.5 except clavulanic acid for TEM-1.

Anti-Bacterial Agents↗

PubMed1995

[Left thoracotomy approach for coronary reoperation].

We experienced reoperative coronary revascularization through a left thoracotomy in 12 patients between June 1992 and June 1994. All patients underwent grafting to the left coronary system except one, who underwent bypass procedure to the atrioventricular branch of the right coronary artery in addition to grafting to the left anterior descending artery, using the pedicled left internal thoracic artery, the gastroepiploic artery as a free graft, or a reversed saphenous vein graft. In one patient, revascularization was accomplished during temporary occlusion of the coronary artery without cardiopulmonary bypass. On the other hand, in 11 patients, cardiopulmonary bypass was used. Coronary bypass procedure was employed under the ventricular fibrillation with hypothermia in those but one, who underwent revascularization under the beating heart with cardiopulmonary support. All patients were hemodynamically stable in postoperative stage, and artificial ventilation time was not prolonged, with a mean time of 15.7 hours. Postoperative morbidity included reexploration for hemorrhage in one patient, convulsion in one, worsening of hemianopsia in one, ventricular arrhythmia in one, and wound complication in one, however, there were no hospital deaths. All patients underwent repeat coronary angiography, which revealed that all grafts were widely patient except one, which had inadequate flow due to diffuse narrowing. Based on these clinical results, we conclude that a left thoracotomy is a useful approach for reoperative coronary bypass procedures to reduce the surgical risk associated with a sternal reentry in properly selected patients.

Adult↗

PubMed1995

[Clinical evaluation of latex photometric immunoassay for serum CA19-9].

Latex photometric immunoassay for CA19-9 was clinically evaluated. Reference interval of the assay was determined as 37U/ml based on the data of 350 healthy donors. CA19-9 values were higher in females than in males (p < 0.01) and distribution of CA19-9 values varied significantly depending on the Lewis blood types of the subjects; namely, subjects with Lea+b- gave highest CA19-9 values followed by those with Lea-b+ and then those with Lea-b- (p < 0.001). In the patients with various malignant diseases, positive rates were high (> 60%) in the patients with pancreatic, hepatic, and colon cancer. Meanwhile, false positive rates were around 25-50% in the patients with pancreatitis, cholelithiasis, benign hepatic disorders and benign lung diseases. When cut-off value was set at 100U/ml, false positive rates decreased drastically, while true positive rates remained slightly decreased. These findings indicate that, in this latex assay, CA19-9 values of 100U/ml or higher should be regarded as high risk factor of malignancy and values in the range of > or = 37U/ml and < 100U/ml might be considered as warning signal of various benign and malignant diseases.

Biomarkers, Tumor↗

PubMed1995

[Multiple systemic embolization from floating thrombus in left atrium].

A 66-year-old woman was referred to our hospital because of cerebral infarction, myocardial infarction and renal infarction. Further examination indicated that she was suffering from mitral valve stenosis with a floating thrombus in the left atrium. She underwent emergency mitral valve replacement and thrombectomy. The thrombus was attached to the left atrium by only four thin and weak strings and removed easily. We think that the thrombus was the precursor of a free-floating ball thrombus without stalk.

Aged↗

PubMed1995

Central cord syndrome secondary to hyperflexion injury of the cervical spine in a child.

A 10-year-old girl sustained hyperflexion injury to the midcervical spine, i.e., wedge compression fractures of the C3, C4, and C5 vertebral bodies, when hit by an automobile, subsequently manifesting central cord syndrome below the C4 spinal cord level. Serial magnetic resonance imaging helped identify the spinal cord lesion at the corresponding levels. Conservative management combined with corticosteroid administration resulted in nearly full neurologic recovery approximately 3 months after the trauma, although a cystic lesion remained present within the spinal cord 11 months after trauma.

Cervical Vertebrae↗

PubMed1994

Characterization of a plasmid-borne and constitutively expressed blaMOX-1 gene encoding AmpC-type beta-lactamase.

A 1954-bp DNA fragment containing the blaMOX-1 gene, identified on a large resident plasmid (pRMOX-1) of Klebsiella pneumoniae NU2936, was sequenced and an open reading frame (ORF) coding for a 390-amino-acid (aa) MOX-1 was found. The total deduced aa sequence of MOX-1 shared considerable homology with that of AmpC-type class C beta-lactamases of Gram- bacteria, especially of Pseudomonas aeruginosa PAO1 [51.3%; 63.8% at the nucleotide (nt) level]. However, the regulatory gene ampR and a 38-bp AmpR-binding region were not present upstream from blaMOX-1, although the expression of P. aeruginosa ampC is directly regulated by AmpR. Possible -35 and -10 regions, a Shine-Dalgarno (SD) sequence and terminators were identified which are peculiar to blaMOX-1. On the other hand, a sequence highly homologous (91.6%) to the region upstream from dhfrX in the In7 integron carried by plasmid pDGO100 was found upstream from blaMOX-1 at nt 1 to 488. No significant difference was detected between the promoter activities of blaMOX-1 in ampD- and ampD+ strains of Enterobacter cloacae, as measured by the chloramphenicol acetyltransferase (CAT) assay. These results clearly show that blaMOX-1 belongs to the group of ampC-related bla genes and that it is expressed constitutively, independently of transcriptional regulators such as AmpR, AmpG and AmpD. Homology analysis among AmpC enzymes or ampC genes implied that integration of the chromosomal ampC gene into a large resident plasmid, followed by transconjugation, was involved in the evolution of blaMOX-1.

Amino Acid Sequence↗

PubMed1994

Gastroepiploic artery graft for anterior descending coronary artery bypass.

In 308 right gastroepiploic artery (GEA) grafting procedures performed for myocardial revascularization, 38 GEA, 34 in situ, and four free grafts were used to bypass the left anterior descending coronary artery (LAD). Indications for using the GEA for the purpose of LAD bypass were: unavailability of the internal thoracic artery (ITA) at reoperation, surgical damage to the ITA at the time of the operation, or an apparently better free flow versus that in the left ITA, particularly in patients with diabetes mellitus in whom it was considered inadvisable to use bilateral ITAs. There were 21 male and 17 female patients with a mean age of 62 years (range, 31 to 77 years). Ten patients had undergone a previous myocardial revascularization. The mean number of distal anastomoses was 2.8 (range, 1 to 5). Concomitantly used conduits were the ITA in 27 patients, saphenous veins in 21 patients, the inferior epigastric artery in 4 patients, and the bovine internal thoracic artery in 1 patient. All but 1 patient survived. Follow-up ranged from 3 to 84 months (mean, 27 months). Postoperative angiography was performed in 33 patients. At the short-term evaluation (mean, 1 month), 32 of 33 (97%) GEA grafts were found to be patent; all 4 GEA grafts studied at the long-term evaluation (mean, 25 months) were also found to be patent. In no patients did angina recur postoperatively. In 25 patients who underwent an exercise study postoperatively, the stress test results were negative in 23.(ABSTRACT TRUNCATED AT 250 WORDS)

Abdominal Muscles↗

PubMed1994

Purification and characterization of dihydrofolate reductase of Plasmodium falciparum expressed by a synthetic gene in Escherichia coli.

We have expressed the dihydrofolate reductase (DHFR) part of the DHFR-thymidylate synthetase complex of P. falciparum in Escherichia coli, by constructing a gene with synthetic oligonucleotides that changed the gene's codon usages. The induced expression in an E. coli cell of the synthetic gene yielded a product that constituted about 30% of the total bacterial protein. The product was precipitated in an inclusion body in a cell. Its enzymatic activity was restored after denaturation and renaturation procedures with guanidine-HCl. Recombinant DHFRs with Ser or Thr at position 108 were prepared. Kinetic characterization showed that the DHFRSer108 has less of an affinity for NADPH and dihydrofolate than the DHFRThr108.

Amino Acid Sequence↗

PubMed1994

Molecular characterization of an enterobacterial metallo beta-lactamase found in a clinical isolate of Serratia marcescens that shows imipenem resistance.

A clinical isolate of Serratia marcescens (TN9106) produced a metallo beta-lactamase (IMP-1) which conferred resistance to imipenem and broad-spectrum beta-lactams. The blaIMP gene providing imipenem resistance was cloned and expressed in Escherichia coli HB101. The IMP-1 was purified from E. coli HB101 that harbors pSMBNU24 carrying blaIMP, and its apparent molecular mass was calculated to be about 30 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Kinetic studies of IMP-1 against various beta-lactams revealed that this enzyme hydrolyzes not only various broad-spectrum beta-lactams but also carbapenems. However, aztreonam was relatively stable against IMP-1. Although clavulanate or cloxacillin failed to inhibit IMP-1, Hg2+, Fe2+, or Cu2+ blocked the enzyme's activity. Moreover, the presence of EDTA in the reaction buffer resulted in a decrease in the enzyme's activity. Carbapenem resistance was not transferred from S. marcescens TN9106 to E. coli CSH2 by conjugation. A hybridization study confirmed that blaIMP was encoded on the chromosome of S. marcescens TN9106. By nucleotide sequencing analysis, blaIMP was found to encode a protein of 246 amino acid residues and was shown to have considerable homology to the metallo beta-lactamase genes of Bacillus cereus, Bacteroides fragilis, and Aeromonas hydrophila. The G+C content of blaIMP was 39.4%. Four consensus amino acid residues, His-95, His-97, Cys-176, and His-215, which form putative zinc ligands, were conserved in the deduced amino acid sequence of IMP-1. By determination of the amino acid sequence at the N terminus of purified mature IMP-1, 18 amino acid residues were found to be processed from the N terminus of the premature enzyme as a signal peptide. These results clearly show that IMP-1 is an enterobacterial metallo beta-lactamase, of which the primary structure has been completely determined, that confers resistance to carbapenems and other broad-spectrum beta-lactams.

Amino Acid Sequence↗

PubMed1994

[A case of coronary artery bypass grafting with coronary-coronary bypass for familial hypercholesterolemia].

Coronary artery bypass grafting in a 68-year-old woman with familial hypercholesterolemia and severe calcification of the ascending aorta was successfully performed. The left internal thoracic artery bypassed the left anterior descending artery, the right gastroepiploic artery bypassed the left circumflex artery and the saphenous vein graft bypassed the right coronary artery, the proximal anastomosis was based on segment 1 and the distal anastomosis was on segment 3 of the right coronary artery (coronary-coronary bypass). The operation was performed with aortic no touch technique which consisted of fibrillatory arrest without aortic cross-clamping, and femoral artery perfusion. The post operative angiogram revealed patency of all graft, and she discharged at 20th post operative day. Coronary-coronary bypass was took into consideration in a patient with severe calcification of the ascending aorta and without suitable in situ arterial graft.

Abdominal Muscles↗

PubMed1994

[Coronary artery bypass grafting for active aortitis syndrome with bilateral coronary ostial stenosis].

Coronary artery bypass grafting was performed on a 31-year-old female for treatment of active aortitis syndrome with unstable angina. Preoperative coronary angiography revealed ostial stenosis of coronary arteries. The ascending aorta was intensely inflamed. In the proximal anastomosis, the ascending aorta was sutured with an autogenous pericardial patch and anastomosed with the saphenous vein (SV) to be jointed to left anterior descending (LAD). At the same time, gastroepiploic artery (GEA) was connected to LAD by taking into account a degenerative change in remote stage. Right coronary artery was anastomosed with right internal thoracic artery (RITA). The postoperative course was satisfactory. On graft angiography SV and RITA were adequately patent, but GEA was unsatisfactorily patent because of its competition with SV for patency. This surgical procedure seemed to be an option to be indicated for a patient with unstable angina at an active inflammatory stage.

Adult↗

PubMed1994

[Reoperative coronary revascularization without cardiopulmonary bypass].

We report a case of successful reoperation of coronary revascularization without cardiopulmonary bypass. The patient was a 62-year-old man, who had undergone coronary artery bypass grafting (CABG) to the LAD and CX with two saphenous vein grafts (SVG) for the left main lesion 12 years before. He required reoperation for unstable angina due to progressive ischemic heart disease and the diseased SVG. Preoperative coronary angiogram revealed total occlusion of major 3 branches and the diseased SVG to the LAD. The reoperation was performed without cardiopulmonary bypass through the repeated median sternotomy for revascularization of the LAD and RCA. The left internal thoracic artery and the gastroepiploic artery were anastomosed to the LAD and RCA under the beating heart without any hemodynamic or electrocardiographic deteriorations. The operation was uneventfully finished in 3 hr 40 min. without the use of blood products. Postoperative angiogram showed both new grafts were widely patent, and he was discharged 14 days after the operation without angina. We also performed 4 other cases of reoperative CABG without cardiopulmonary bypass, and conclude that this technique is a safe and effective alternative in a carefully selected group of patients for reoperative CABG to reduce several technical problems related to coronary reoperation.

Abdominal Muscles↗