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Biomedical subjects

H Kanazawa

Publications and source records attributed to H Kanazawa.

At least 271 records · Page 15Linked to original sources
PubMed1985

[The pharmacokinetics and transfer into atrial muscle of ceftizoxime in open heart surgery].

A study of serum and atrial muscle concentration of ceftizoxime (CZX) was performed in 28 adult patients having cardiac surgery. Patients were administrated a single dose of CZX 2 g intravenously. Serum and atrial muscle samples were obtained at suitable intervals before extracorporeal circulation, and assayed CZX concentration. Serum concentrations were 54.5 micrograms/ml at 60 minutes, and 21.8 micrograms/ml at 180 minutes. Atrial tissue levels were 29.9 micrograms/g at 60 minutes, and 14.8 micrograms/g at 180 minutes. Tissue/serum concentration ratio was high value; 0.549 at 60 minutes, 0.679 at 180 minutes. Judging from its favorable transfer into atrial muscle and MIC80 against the suspected pathogen, CZX seemed to be clinically useful in the treatment of infection after cardiac surgery.

Adolescent↗
PubMed1985

[Radiotherapy of brain metastases of breast cancer].

Twenty-two patients with brain metastases of breast cancer treated by irradiation between 1974 and 1983 were reviewed. Significant neurologic improvement was obtained in 80%. The median duration of symptomatic palliation caused by irradiation and the median survival from the diagnosis of brain metastases were 3.0 months and 5.0 months, respectively. As a favorable prognosis can be expected for patients showing good performance, minor neurologic disorders, a long latent period between the first treatment of the primary lesion and the onset of the brain metastasis, and without extracranial metastases, irradiation with more than 40 Gy is recommended. Sequential CT examinations were useful to judge the effectiveness of treatment.

Adult↗
PubMed1984

A phenylalanine for serine substitution in the beta subunit of Escherichia coli F1-ATPase affects dependence of its activity on divalent cations.

A mutant (KF11) of Escherichia coli H+-translocating ATPase (F1-F0) has a single point mutation in the beta subunit of F1 that has lost 90% of its Mg2+-dependent ATPase activity (Kanazawa, H., Horiuchi, Y., Takagi, M., Ishino, Y., and Futai, M. (1980) J. Biochem. (Tokyo) 88, 695-703). The mutation was mapped at about the 500th nucleotide residue from the 5' end of the beta subunit gene by a genetic recombination test on the physical map of the cistron coding for the beta subunit. The mutant allele of KF11 (uncD11) was cloned on a hybrid plasmid (pKF11) via DNA isolated from a lambda uncD11 transducing phage. Restriction fragments of pKF11 containing the estimated mutation site were subjected to polyacrylamide gel electrophoresis under conditions where strands were separated into single strands. The two strands of a DNA segment, which was shown to carry an altered base, showed anomalous migration compared with those from the wild-type fragment. The results confirmed the result of mapping of the altered site by genetic tests. On the basis of these results, the nucleotide sequence of the mutated gene was determined, and a single base change of the 524th cytosine to thymine resulting in a phenylalanine for serine substitution at residue 174 of the beta subunit was found. This result, together with results on the altered properties of F1 from KF11 reported previously, indicates that residue 174 is essential for the Mg2+-dependent ATPase activity of F1 but not for the Ca2+-dependent ATPase activity.

Amino Acid Sequence↗
PubMed1984

Replacement of serine 373 by phenylalanine in the alpha subunit of Escherichia coli F1-ATPase results in loss of steady-state catalysis by the enzyme.

The mutant allele (uncA401) of the gene for the alpha subunit of Escherichia coli F1-ATPase was cloned from the total DNA of the mutant AN120 on a hybrid plasmid pAN120. Determination of the DNA sequence of the alpha subunit gene from pAN120 revealed a single base change of cytosine at nucleotide residue 1118 to thymine and indicated that serine 373 was replaced by phenylalanine. It has been reported that the mutant F1 is defective in a step of steady-state catalysis, whereas its single turnover process is normal (Kanazawa, H., Noumi, T., Matsuoka, I., Hirata T., and Futai, M. (1984) Arch. Biochem. Biophys. 228, 258-269). Thus, we concluded that serine 373 in the alpha subunit is essential for steady-state catalysis by F1-ATPase.

Alleles↗
PubMed1984

Detection of ATP-dependent conformational change in the F1 portion and beta subunit of Escherichia coli H+-ATPase using 8-anilinonaphthalene-1-sulfonate.

The bindings of ATP to Escherichia coli coupling factor ATPase (F1) and its isolated beta subunit were studied with 8-anilinonaphthalene-1-sulfonate (ANS). The fluorescence of ANS increased upon addition of F1 or the beta subunit, and this fluorescence was quenched on addition of ATP. Thus, ANS bound to the hydrophobic region of F1 or the beta subunit, and the binding of ATP was detected as a quenching of ANS fluorescence. The quenching of the fluorescence suggested that F1 or the beta subunit underwent a conformational change on binding to ATP. With and without ATP, 5.0 and 3.6 mol of ANS, respectively, bound to 1 mol of F1. On the other hand, with or without ATP about 1 mol of ANS bound to beta. The fluorescence quenching was specific for ATP and was not observed with GTP or CTP. It was dependent on pH, being higher at acidic pH, but it was not enhanced by MgCl2. The dissociation constants of F1 and the beta subunit for ATP were estimated to be 10(-4)-10(-5) M. The significance of these binding sites is discussed in relation to the mechanism of the ATPase.

Escherichia coli↗
PubMed1984

Nucleotide sequence of the melB gene and characteristics of deduced amino acid sequence of the melibiose carrier in Escherichia coli.

The nucleotide sequence of the melB gene coding for the melibiose carrier in Escherichia coli has been determined. The melibiose carrier is predicted to consist of 469 amino acid residues, resulting in a protein with a molecular weight of 52,029. The predicted carrier protein is highly hydrophobic (70% nonpolar amino acid residues). The hydropathic profile suggests that there are 10 long hydrophobic segments in the primary structure of the carrier protein. Most of them seem to traverse the membrane. Although the hydropathic profile of the melibiose carrier is similar to that of the lactose carrier as a whole, homology in the primary structure between the two carriers is very low. Furthermore, no homology in the nucleotide sequence is found in the structural genes for the two carriers. However, the nucleotide sequences of the intergenic regions are very similar between the melibiose operon and the lactose operon. There is a typical intercistronic regulatory sequence in the 3'-flanking region of the melB as well as in that of the lacY, which suggests the presence of another gene downstream of the melB.

Amino Acid Sequence↗
PubMed1984

Comparison of F1's of oxidative phosphorylation from Escherichia coli and Salmonella typhimurium and demonstration of interchangeability of their subunits.

The peripheral membrane portion (SF1) of proton-translocating ATPase of Salmonella typhimurium and its alpha, beta, and gamma subunits were purified and compared with the same portion (EF1) from Escherichia coli. The alpha, beta, and gamma subunits of these F1's were found to be mutually interchangeable, and all possible combinations of the three subunits from EF1 and SF1 showed ATPase activity. Both F1's could bind functionally to the integral membrane part (F0) of either bacterium, suggesting that F0 and F1 are interchangeable in these two bacteria and thus that the two F1's are closely similar at the level of subunit structure. However, SF1 differed from EF1 in some enzymological properties such as its specific activity and susceptibilities to sodium dodecyl sulfate and methanol. The specific ATPase activity of EF1 was more than twice that of SF1, and hybrid enzymes containing the beta subunit of EF1 had higher activity than other hybrids. Amino acid analysis suggested that the primary structures of the alpha subunits of the two F1's are less homologous than those of the beta subunits. Thus, the primary structure of the alpha subunit may be more species specific than that of the beta subunit.

Amino Acids↗
PubMed1984

Physical and genetic characterization of the melibiose operon and identification of the gene products in Escherichia coli.

We have identified hybrid plasmids carrying the melibiose operon of Escherichia coli in a colony bank of Clarke and Carbon (Tsuchiya, T., Ottina, K., Moriyama, Y., Newman, M., and Wilson, T. H. (1982) J. Biol. Chem. 257, 5125-5128). Using one of the plasmids as a starting material, the DNA fragments containing the melibiose operon were recloned in a vector pBR322. Restriction maps were prepared, and several DNA segments were subcloned into pBR322. Genetic complementation tests and recombination analyses using those plasmids and melA- and melB- mutants as well as biochemical analyses of mel mutants transformed with those plasmids enabled us to determine the physical location of promoter, melA, and melB on the DNA segment. The size of the melAB region was about 3,000 base pairs. Gene products were identified using maxicells harboring plasmids carrying the melibiose operon. The apparent molecular weight of the alpha-galactosidase (coded by melA) was about 50,000 and that of the melibiose carrier (coded by melB) was about 31,000, as estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The melibiose carrier was also identified as a 30,000-dalton protein in reconstituted proteoliposomes which possessed melibiose transport activity.

Disaccharides↗
PubMed1984

Insertions of transposable elements in the promoter proximal region of the gene cluster for Escherichia coli H+-ATPase: 8 base pair repeat generated by insertion of IS1.

A plasmid pKY159 (Yamaguchi and Yamaguchi 1983) carrying a promoter proximal portion of the gene cluster of the proton-translocating ATPase (H+-ATPase) of Escherichia coli causes growth inhibition of wild-type cells. Insertion of a transposable element in this plasmid released this inhibitory effect. In analyzing this inhibitory effect, we determined the insertion points at the nucleotide-sequence level of transposable elements on 30 independent derivatives of pKY159 . Insertions of IS1, IS5, and gamma delta were found between the promoter and the gene for a possible component of 14,000 daltons of the H+-ATPase. Of 31 insertions, 26 were of IS1 and were located at the same site, indicating that this site is a hotspot for IS1 insertion and that IS1 insertion is much more frequent than that of IS5 or gamma delta in this region. Four different sites for IS1 insertion were found; in two of these an 8 base pair (bp) duplicate of the target sequence ( AAAAACGT and AAACGTTG ) was generated, while in the other two a 9 bp duplicate was found. In all cases in this study the nucleotide sequence of IS1 was the same as that of IS1-K. In the two cases with an 8 bp duplicate in different sites, a common 6 bp sequence ( AAACGT ) was found. These results suggested that generation of the 8 bp duplicate is related to the common sequence rather than a mutation in IS1 suggested by Iida et al. (1981) and also suggested that the essential length of the duplicate is 8 bp or less than 8 bp. A 6 bp sequence ( GTGATG ) homologous to the end portion of IS1 was found at the hotspot , but not at other sites, suggesting that this homology contributed to the high frequency of IS1 insertion.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1984

F1ATPase of Escherichia coli: a mutation (uncA401) located in the middle of the alpha subunit affects the conformation essential for F1 activity.

F1ATPase from the Escherichia coli mutant of H+-ATPase, AN120 (uncA401), has less than 1% of the wild type activity and has been shown to be defective in the alpha subunit by in vitro reconstitution experiments. In the present study, the mutation site was located within a domain of the subunit by recombinant DNA technology. For this, a series of recombinant plasmids carrying various portions of the alpha subunit gene were constructed and used for genetic recombination with AN120. Analysis of the recombinants indicated that the mutation site could be located between amino acid residues 370 and 387. The biochemical properties of the mutant F1 were analyzed further using the fluorescent ATP analog DNS-ATP (2'-(5-dimethylaminonaphthalene-1-sulfonyl)-amino-2'-deoxy ATP). The single turnover process of E. coli F1ATPase proposed by Matsuoka et al. [(1982) J. Biochem. 92, 1383-1398.] was compared in the mutant and wild type F1's. Mutant F1 bound DNS-ATP and hydrolyzed it as efficiently as wild type F1. Results showed that binding of ATP to a low affinity site, possibly in the beta subunit, caused decrease of fluorescence of DNS-ATP in the wild type F1 and that this effect of ATP binding was inhibited by DCCD (dicyclohexyl carbodiimide). However, this effect was not inhibited by DCCD in the mutant F1, suggesting that in the proposed process some step(s) after ATP binding to the low affinity site differed in the mutant and wild F1's. When Pi was added to F1 bound to DNS-ATP or to aurovertin, a fluorescent probe capable of binding to the beta subunit, the opposite changes of fluorescence of these probes in the mutant and wild type F1's were observed, suggesting that the conformational change induced by phosphate binding was altered in the mutant F1. On the basis of the estimated mutation site and the biochemical properties of the mutant F1, the correlation of the domain of this site in the alpha subunit with the function of F1 ATPase is discussed.

Adenosine Triphosphate↗
PubMed1984

Overproduction of subunit a of the F0 component of proton-translocating ATPase inhibits growth of Escherichia coli cells.

A hybrid plasmid, pKY159, carrying the promoter and the proximal region of the gene cluster for proton-translocating ATPase caused growth inhibition of Escherichia coli cells (K. Yamaguchi and M. Yamaguchi, J. Bacteriol. 153:550-554, 1983). The mechanism of this growth inhibition was studied, especially in terms of the responsible gene(s). Insertion of IS1, IS5, or gamma delta between the promoter and the gene for a possible component of the ATPase of 14,000 daltons (14K protein) released the inhibitory effect by pKY159. Deletion of the gene for subunit a also released the effect. However, deletion in the gene for the 14K protein released the effect only with an additional insertion within the gene. These results suggested that overproduction of subunit a is closely related to growth inhibition, whereas the 14K protein is not.

Chromosome Deletion↗
PubMed1984

Monoclonal antihuman thyroglobulin antibodies.

Six murine hybridomas secreting monoclonal antihuman thyroglobulin (Tg) antibodies (TAK 1-6) were established by cell fusion techniques. Solid phase RIA was employed to detect the anti-Tg antibody in culture supernatants of hybridomas. The characteristics of these monoclonal antibodies were analyzed by radioimmune blocking assay using rat Tg, human glycoproteins, thyroid hormones, and various preparations of Tg obtained from patients with thyroid disease as inhibitors. In the same system, competitive inhibition studies between 125I-labeled and unlabeled monoclonal antibodies were carried out to determine whether these antibodies recognized the same antigenic determinant. TAK 2 and 3 reacted with human Tg specifically and had equal binding activity using various preparations of human Tg. The other four monoclonal antibodies (TAK 1, 4, 5, and 6) cross-reacted with xenogeneic Tg (rat Tg) and their affinity for human Tg increased as the iodine content of Tg increased. Tg binding to TAK 1 and 4 was inhibited by T4, whereas Tg binding to TAK 5 and 6 was not inhibited by any thyroid hormone or their precursors. In conclusion, we prepared six monoclonal antihuman Tg antibodies. One group is specific for human Tg and recognizes the framework structure unmodified by iodination; the second group reacts with iodination-related epitopes other than iodoamino acids, and the third group recognizes determinants consisting of T4. These monoclonal antibodies provide important probes to detect the polymorphism of human Tg.

Animals↗
PubMed1984

Hereditary deficiency of OKT4-positive cells: studies for mode of inheritance and lymphocyte functions.

Two Graves' patients were found to have no OKT4+ cells in their peripheral blood lymphocytes (PBL). However, the reactivity of their lymphocytes with T4(T4A) and anti Leu-3a was normal and no autoantibodies to the OKT4 determinant were found in their sera. Cytofluorographic analysis of PBL from their family members showed three types of immunofluorescence profiles with OKT4. The first type was the complete OKT4+ cell deficiency, the second was the normal percentage of OKT4+ cells with half immunofluorescence intensity and the third was the normal staining pattern with OKT4. Phytohaemagglutinin (PHA) and pokeweed mitogen (PWM) induced blastogenesis; PWM induced IgG synthesis, autologous and allogeneic mixed lymphocyte reaction, and Interleukin-2 (IL-2) production of their PBL were also normal. These results suggest that (i) the expression of determinant to OKT4 is transmitted as autosomal incomplete dominant trait and (ii) OKT4+ cell deficiency is not accompanied by a lack of the inducer/helper subset of T cells.

Antibodies, Monoclonal↗