PubMed Health⌕ Search

Biomedical subjects

T Miyata

Publications and source records attributed to T Miyata.

At least 685 records · Page 38Linked to original sources
PubMed1984

Structure and expression of the alkA gene of Escherichia coli involved in adaptive response to alkylating agents.

Nucleotide sequence of a DNA fragment containing the alkA gene and its control region has been determined using a chemical method. Only one open reading frame responsible for 3-methyladenine DNA glycosylase II was found. The hypothetical polypeptide deduced from the DNA sequence, with a molecular weight of 31,400, has an amino-terminal sequence and total amino acid composition identical to that of purified 3-methyladenine DNA glycosylase II. We constructed hybrid plasmids carrying an alkA'-lacZ' fusion, with the proper control region for alkA expression. A hybrid polypeptide with beta-galactosidase activity was formed when lac mutant cells harboring such plasmids were incubated with low doses of N-methyl-N'-nitro-N-nitrosoguanidine or methylmethane sulfonate. Other DNA-damaging agents, such as ethylmethane sulfonate, nalidixic acid, and ultraviolet light did not induce the enzyme activity. The induction was controlled by the ada and adc, but not by the recA and lexA genes.

Alkylating Agents↗
PubMed1984

Quantification by selected ion monitoring of pipecolic acid, proline, gamma-aminobutyric acid and glycine in rat brain.

A procedure for the simultaneous analysis of brain pipecolic acid, proline, gamma-aminobutyric acid and glycine--amino acids with potent inhibitory actions on the central nervous system--was developed. The identification and quantification of the amino acids were performed with a gas chromatographic--mass spectrometric--computer system using deuterium-labelled amino acids as the internal standards. After separation of the amino acids by high-performance liquid chromatography, the methyl ester heptafluorobutyryl derivatives were prepared. The lower limit of quantification for this method is at the picomole level. The usefulness of this chromatographic procedure has been demonstrated by measurement of trace amounts of pipecolic acid in rat brain.

Animals↗
PubMed1984

Amino acid sequence of the coagulogen from Limulus polyphemus hemocytes.

The complete amino acid sequence of the coagulogen from hemocyte lysates of Limulus polyphemus has been determined by sequencing the peptides obtained from tryptic, chymotryptic, staphylococcal protease V8 and lysyl endopeptidase digestions. These results established the following sequence: (formula; see text) Limulus coagulogen consists of a single chain with a total of 175 amino acid residues and the molecular weight is calculated to be 19,675. It contains 16 half-cystines in disulfide linkages, with 5 half-cystines located in a cluster in the COOH-terminal 14 residues. The sequence of Limulus coagulogen is very close to that for the coagulogen of Tachypleus tridentatus (Japanese horseshoe crab), having 69% sequence homology. The 16 half-cystines of these coagulogens are in the same positions, suggesting a very similar conformation. Moreover, the COOH-terminal tripeptide regions of the A chain (from the NH2-terminal end to Arg-18) and peptide C (from Lys-19 to Arg-46), both of which seem to interact with a Limulus clotting enzyme to liberate peptide C, are completely conserved. From secondary structure predictions by the method of Chou and Fasman (Chow, P.Y., and Fasman, G. D. (1974) Biochemistry 13, 211-222), the coagulogen appears to contain an alpha-helical region in the peptide C segment, released by the clotting enzyme, suggesting a marked conformational change in the transformation of the coagulogen to the coagulin gel. beta-sheet and reverse turn regions are distributed in the B chain segment (from Gly-47 to the COOH-terminal end). It is likely that the 16 half-cystines and abundant beta-sheet structure make the coagulogen molecule compact.

Amino Acid Sequence↗
PubMed1984

Influence of cerebral cortex stimulation upon cough-like spasmodic expiratory response (SER) and cough in the cat.

In 27 pentobarbitalized cats, the influence of electrical stimulation of the cerebral cortex upon the spasmodic expiratory response (SER) was studied and compared with cortical influences on coughing induced by stimulation of the superior laryngeal nerve (sup. laryngeal N.). This cortical influence was evoked by electrical stimulation of the cortical nucleus of amygdala (Aco), and was very similar to coughing accompanying changes in emotional behavior and was depressed more effectively by psychotropics than by centrally acting antitussives like codeine. When anterior cingulate gyrus (ant. cingulate G.) or orbital gyrus (orbital G.) were stimulated simultaneously with Aco or sup. laryngeal N., weak stimulation was sufficient to inhibit SER, while stronger stimuli were needed for the suppression of cough. If the same cortical regions were stimulated after initiation of SER or cough, SER was markedly suppressed but cough little affected. Production of SER was facilitated by simultaneous stimulation of the piriform lobe (piriform L.) or olfactory tract (olfactory T.), whereas cough production was facilitated by simultaneous stimulation of the suprasylvian gyrus. These results suggest that SER and coughing are differently controlled by the cerebral cortex, and that SER is modulated by the limbic cortex, in particular, by ant. cingulate G., orbital G. and piriform L. The mechanism of modulation for SER is discussed.

Amygdala↗
PubMed1984

Evolution and structure of two ADP-ribosylation enterotoxins, Escherichia coli heat-labile toxin and cholera toxin.

Nucleotide sequence comparisons of the heat-labile enterotoxin (LTh) genes of E. coli pathogenic for humans with cholera toxin (CT) genes suggest that the two toxin genes have evolved from a common ancestry by a series of single base changes, while conserving the catalytic fragment A1 (ADP-ribose transferase). Based on the local hydrophilicity profiles of LTh and CT peptides, a transmembrane segment appears to be present in A1 in both toxins.

Amino Acid Sequence↗
PubMed1984

Concerted evolution of the mouse immunoglobulin gamma chain genes.

The nucleotide sequences of the immunoglobulin heavy-chain constant region genes of mouse, C gamma 3, C gamma 1, C gamma 2b and C gamma 2a, together with that of a human equivalent C gamma 4 were compared. All the six pairs of genes within the mouse C gamma gene family contain DNA segments that exhibit marked homology, whereas no such segmental homology was found in interspecies comparisons. This result indicates that the four C gamma genes of the mouse evolved concertedly by exchanging parts of their genetic information with each other either by gene conversion or by double unequal crossing-over. Another example of such concerted evolution was found in gene regions encoding membrane domains of the mouse C gamma chains. We also searched for such segmental homologies in other mammalian C gamma gene families and found at least two more examples in man and guinea-pig. In the mouse C gamma gene family, the silent positions of an exon encoding the third domain of C gamma chains show much greater divergence in sequence than other regions, indicating that the genetic information encoded by this gene region was least scrambled during recent evolution. A phylogenetic tree constructed from the nucleotide differences of this exon demonstrates that at least two C gamma genes had already existed before mammalian radiation. Based on these results, evolution of mammalian C gamma gene families is discussed.

Amino Acid Sequence↗
PubMed1984

Separation of macrophages from mouse peritoneal exudate cells with substrata coated with chemically modified collagens.

A cell-separation technique was designed on the basis of specific interaction between macrophages and chemically modified collagens in the presence of serum. When unseparated mouse peritoneal exudate cells containing approximately 50% macrophages and 50% lymphocytes were incubated in dishes coated with unmodified and chemically modified collagens, only macrophages adhered more rapidly and in greater numbers to succinylated and methylated collagen than to unmodified collagen. The adherent macrophages could be easily detached from the substrata by divalent cation chelating agents. The purity and recovery of macrophages separated by this method were approximately 92-94% and 41-48%, respectively.

Animals↗
PubMed1984

Evolutionary implications of error amplification in the self-replicating and protein-synthesizing machinery.

Evolutionary constraints operating on animal mitochondrial tRNA were estimated to be reduced to about 1/30 of those that apply to cytoplasmic tRNA. In the nuclear-cytoplasmic system, an effect of a mutation in tRNA is likely to be amplified through positive feedback loops consisting of DNA polymerases, RNA polymerases, ribosomal proteins, aminoacyl-tRNA synthetases, tRNA processing enzymes, and others. This amplification phenomenon is called an "error cascade" and the loops that cause it are called "error loops." The freedom of evolutionary change of cytoplasmic tRNA is expected to be severely restricted to avoid the error cascade. In fact, cytoplasmic tRNA is highly conserved during evolution. On the other hand, in the animal mitochondrial system, all of the proteins involved in error loops are coded for in the nuclear genome and imported from the cytoplasm, and accordingly the system is free from the error cascade. The difference in constraints operating on animal tRNA between cytoplasm and mitochondria is attributed to the presence or absence of error loops. It is shown that the constraints on mitochondrial tRNA in fungi are not as relaxed as those in animals. This observation is attributed to the presence of an error loop in fungal mitochondria, since at least one protein of the mitochondrial ribosome is coded for in the mitochondrial genome of fungi. The evolutionary rates of proteins involved in the processing of genetic information are discussed in relation to the error cascade.

Animals↗
PubMed1984

Purification and characterization of four forms of cytochrome P-450 from liver microsomes of phenobarbital-treated and 3-methylcholanthrene-treated rats.

Four forms of cytochrome P-450, tentatively designated PB-1, PB-2, MC-1, and MC-2, were purified from liver microsomes of rats treated with phenobarbital (PB-1 and PB-2) or 3-methylcholanthrene (MC-1 and MC-2). Each purified form showed a single protein-staining band on SDS-polyacrylamide gel electrophoresis giving a minimum molecular weight of 56,000 (MC-1), 53,000 (PB-1), 53,000 (MC-2), or 49,000 (PB-2). PB-1 and MC-1 were the major cytochrome P-450 components inducible by phenobarbital (PB) and 3-methylcholanthrene (MC), respectively. Antibodies prepared against each form of purified cytochrome P-450 did not cross-react with heterologous antigens in Ouchterlony double diffusion tests, confirming the immunological distinctness of the four forms. The CO-compounds of reduced PB-1 and PB-2 had an absorption maximum at 450 nm, whereas those of MC-1 and MC-2 had a maximum at 447 nm. Judging from the oxidized absolute spectra, MC-2 was of high spin type and the others were of low spin type. Amino acid analysis revealed considerable differences among the purified four forms of cytochrome P-450, and the amino acid sequences of their NH2-terminal portions confirmed that the four forms were different proteins. In a reconstituted system containing NADPH and NADPH-cytochrome P-450 reductase, PB-1 and PB-2 oxidized benzphetamine at high rates, but their oxidation of benzo(a)pyrene was much slower than that by MC-1, which catalyzed rapid hydroxylation of benzo(a)pyrene but had low activity with benzphetamine. The quantity of each form of cytochrome P-450 in microsomes was determined by quantitative immunoprecipitation, and selective induction of PB-1 and MC-1 by PB and MC, respectively, was confirmed. Some induction of PB-2 and MC-2 by the corresponding inducers was also noticed. PB group P-450's were not increased by MC treatment, nor were MC group P-450's by PB.

Amino Acids↗
PubMed1984

Induction of a phenobarbital-inducible form of cytochrome P-450 in rat liver microsomes by 1,1-di(p-chlorophenyl)-2,2-dichloroethylene.

When 1,1-di(p-chlorophenyl)-2,2-dichloroethylene (DDE) (100 mg/kg body weight) was injected into rats, the benzphetamine N-demethylation and 7-ethoxycoumarin O-deethylation activities of liver microsomes increased by 7-fold and 3-fold, respectively, at 7 days after the injection, whereas the benzo(a)pyrene 3-hydroxylation activity did not increase. The content of cytochrome P-450 in the microsomes increased 2.5-fold at 7 days. By the use of the antibodies to a phenobarbital (PB)-inducible form (P-450(PB-1)) and a 3-methylcholanthrene (MC)-inducible form (P-450(MC-1)) of cytochrome P-450, the contents of P-450(PB-1) and P-450(MC-1) in the liver microsomes of DDE-treated rats were measured. The form of cytochrome P-450 immunoprecipitable with anti-P-450(PB-1) antibodies increased by 10-fold at 7 days. A major component of cytochrome P-450 in the liver microsomes of DDE-treated rats, which was tentatively designated P-450(DDE), was purified. P-450 (DDE) was compared with P-450(PB-1), and they were found to be indistinguishable by the following criteria: 1) chromatographic behavior on aminooctyl-Sepharose 4B, hydroxyapatite, and DEAE-cellulose columns, 2) minimum molecular weight determined by SDS-polyacrylamide gel electrophoresis, 3) spectral properties, 4) immunoreactivity, 5) amino acid composition, 6) peptide mapping, 7) NH2-terminal amino acid sequence, 8) catalytic activities. We concluded that DDE and PB induce an identical form of P-450 in rat liver microsomes, although DDE is apparently very different from PB in chemical structure.

Amino Acid Sequence↗
PubMed1984

A 26K fragment of troponin T from rabbit skeletal muscle.

A 26K fragment of troponin T, which was produced by endogenous proteases in rabbit skeletal muscle, was isolated by SE-Sephadex column chromatography. This fragment sensitized both superprecipitation and ATPase of actomyosin to calcium ions, to the same extent as troponin T. There was no difference in affinity for tropomyosin between this fragment and troponin T as examined by affinity chromatography. Amino acid analysis showed that this fragment consisted of residues Ala-46-Lys-259 of troponin T. The N-terminal 45 residues of troponin T, therefore, are not essential for the physiological action of troponin T. It was also observed that Ca2+-activated neutral protease digested troponin T into the 26K fragment in the native thin filament, while the protease digested troponin T in a different way in the reconstituted thin filament.

Amino Acids↗
PubMed1984

Amino acid sequence of Japanese horseshoe crab (Tachypleus tridentatus) coagulogen B chain: completion of the coagulogen sequence.

The complete amino acid sequence of the B chain derived from Tachypleus tridentatus coagulogen was determined. It consisted of a total of 129 amino acid residues with a NH2-terminal glycine and COOH-terminal phenylalanine. Sequence studies of the whole B chain and the fragments obtained from the digests with trypsin, alpha-chymotrypsin, thermolysin and Staphylococcal protease V8 showed the following sequence: (sequence; see text) These structural studies of the B chain and the previously established amino acid sequences of the A chain and peptide C derived from T. tridentatus coagulogen, now make it possible to complete the whole sequence of coagulogen consisting of 175 amino acid residues with the molecular weight of 19,723.

Amino Acid Sequence↗
PubMed1984

The amino acid sequence of coagulogen isolated from southeast Asian horseshoe crab, Tachypleus gigas.

The amino acid sequence of coagulogen isolated from Southeast Asian horseshoe crab (Tachypleus gigas) has been determined. The NH2-terminal sequence of the first 51 residues was obtained by automated Edman degradation. The intact protein was then treated with a Tachypleus clotting enzyme, to form a gel and to remove an internal peptide C (28 residues) located near the NH2-terminal portion. The gel protein, which consisted of A chain (18 residues) and B chain (129 residues), was S-alkylated and the resulting two chains were separated by acetone precipitation. Among these segments, A chain and peptide C were assigned to the NH2-terminal portion of whole coagulogen, as judged from their amino acid compositions. On the other hand, the covalent structure of B chain was determined by sequencing the peptides obtained from its tryptic digest. The alignments of the tryptic peptides were deduced from the sequence homology in comparison with the previously established B chain sequence of Japanese horseshoe crab (T. tridentatus) coagulogen. T. gigas coagulogen had a total of 175 amino acids and a calculated molecular weight of 19,770. When the sequence was compared with those of Japanese and American horseshoe crab (Limulus polyphemus) coagulogens, extensive structural homology was found: T. tridentatus/T. gigas, 87% and L. polyphemus/T. gigas, 67%. This comparison suggests that Japanese and Southeast Asian horseshoe crabs have a crab, based on amino acid sequence data.

Amino Acid Sequence↗
PubMed1984

Plasminogens Tochigi II and Nagoya: two additional molecular defects with Ala-600----Thr replacement found in plasmin light chain variants.

Previous studies in our laboratories (Miyata, T., et al. (1982) Proc. Natl. Acad. Sci. U.S. 79, 6132-6136) showed that the structural defect in a hereditarily abnormal plasminogen, plasminogen Tochigi, is due to replacement of Ala by Thr at position 600 from the NH2-terminal end. In the present studies, two abnormal plasminogens, plasminogens Tochigi II and Nagoya, obtained from other family members were analyzed to identify the structural impairment in these molecules. Amino acid sequence analysis of one of the tryptic peptides isolated, respectively, from plasminogens Tochigi II and Nagoya indicated that in both cases, Ala-600 (equivalent to Ala-55 of the chymotrypsin numbering system) had been replaced by Thr. No other substitutions at the active site and substrate-binding site residues, namely, His-57, Asp-102, Ser-195, and Asp-189, were found in the plasmin light chain variants, indicating that all these residues are intact. Moreover, the NH2-terminal heptapeptide sequences of the plasmin light chain variants isolated from plasminogens Tochigi II and Nagoya were identical to the sequence determined for the normal control. These results indicate that the absence of proteolytic activity of both abnormal molecules is due to the same amino acid substitution as that of previously reported plasminogen Tochigi.

Alanine↗
PubMed1984

Amino acid sequence of NADH-cytochrome b5 reductase of human erythrocytes.

The amino acid sequence of soluble NADH-cytochrome b5 reductase purified from normal human erythrocytes was determined as one approach to understand the hereditary disease of a deficiency of this enzyme. The protein is hydrophilic as a whole, but two regions, from Phe-36 to Ile-71 and from Met-231 to Phe-275, were found to be highly hydrophobic. The sequence of the latter region is particularly unique, and rich in proline (20%). The sequence of the amino-terminal region was very similar to the partial sequences of the corresponding regions of the enzymes from pig and steer liver microsomes.

Amino Acid Sequence↗