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Biomedical subjects

H Toh

Publications and source records attributed to H Toh.

At least 55 records · Page 3Linked to original sources
PubMed1997

Changes in the growth and enzyme level of Zymomonas mobilis under oxygen-limited conditions at low glucose concentration.

Zymomonas mobilis growing aerobically with 20 g glucose-1 (carbon-limited) in a chemostat exhibited an increase in both the molar growth yield (Yx/s) and the maximum molar growth yield (Yx/smax) and a decrease in both the specific substrate consumption rate (qs) and the maintenance energy consumption rate (me). Stepwise increase in the input oxygen partial pressure showed that anaerobic-to-aerobic transitional adaptation occurred in four stages: anaerobic (0 mm HgO2), oxygen-limited (7.6- 230 mm HgO2), intermediate (273 mm HgO2), and oxygen excess (290 mm HgO2). The steady-state biomass concentration, Yx/s, and intracellular ATP content increased between oxygen partial pressures of 7.6 and 120 mm HgO2, accompanied by a decrease in the qs and the specific acid production rate. The membrane ATPase activity decreased with increasing oxygen partial pressure and reached its lowest levels at 273 mm HgO2, which was the highest input oxygen partial pressure where steady-state conditions were possible. Glucokinase, glucose-6-phosphate dehydrogenase, glyceraldehyde-3-phosphate dehydrogenase, and alcohol dehydrogenase activities also decreased when the oxygen partial pressure was increased above 15 mm Hg, whereas pyruvate decarboxylase was unaffected by aeration. Growth inhibition at 290 mm HgO2 was characterised by a drastic reduction in the pyruvate kinase activity and a collapse in the intracellular ATP pool. The growth and enzyme data suggest that at low glucose concentrations and oxygen-limited conditions, the increase in biomass yields is a reflection of a redirection of ATP usage rather than a net increase in energy production.

Adenosine Triphosphatases↗
PubMed1997

Introduction of a distance cut-off into structural alignment by the double dynamic programming algorithm.

Two approximations were introduced into the double dynamic programming algorithm, in order to reduce the computational time for structural alignment. One of them was the so-called distance cut-off, which approximately describes the structural environment of each residue by its local environment. In the approximation, a sphere with a given radius is placed at the center of the side chain of each residue. The local environment of a residue is constituted only by the residues with side chain centers that are present within the sphere, which is expressed by a set of center-to-center distances from the side chain of the residue to those of all the other constituent residues. The residues outside the sphere are neglected from the local environment. Another approximation is associated with the distance cut-off, which is referred to here as the delta N cut-off. If two local environments are similar to each other, the numbers of residues constituting the environments are expected to be similar. The delta N cut-off was introduced based on the idea. If the difference between the numbers of the constituent residues of two local environments is greater than a given threshold value, delta N, the evaluation of the similarity between the local environments is skipped. The introduction of the two approximations dramatically reduced the computational time for structural alignment by the double dynamic programming algorithm. However, the approximations also decreased the accuracy of the alignment. To improve the accuracy with the approximations, a program with a two-step alignment algorithm was constructed. At first, an alignment was roughly constructed with the approximations. Then, the epsilon-suboptimal region for the alignment was determined. Finally, the double dynamic programming algorithm with full structural environments was applied to the residue pairs within the epsilon-suboptimal region to produce an improved alignment.

Algorithms↗
PubMed1996

Similarity among the Drosophila (6-4)photolyase, a human photolyase homolog, and the DNA photolyase-blue-light photoreceptor family.

Ultraviolet light (UV)-induced DNA damage can be repaired by DNA photolyase in a light-dependent manner. Two types of photolyase are known, one specific for cyclobutane pyrimidine dimers (CPD photolyase) and another specific for pyrimidine (6-4) pyrimidone photoproducts[(6-4)photolyase]. In contrast to the CPD photolyase, which has been detected in a wide variety of organisms, the (6-4)photolyase has been found only in Drosophila melanogaster. In the present study a gene encoding the Drosophila(6-4)photolyase ws cloned, and the deduced amino acid sequence of the product was found to be similar to the CPD photolyase and to the blue-light photoreceptor of plants. A homolog of the Drosophila (6-4)photolyase gene was also cloned from human cells.

Amino Acid Sequence↗
PubMed1996

In vitro analysis of peritoneal adhesions in peritonitis.

Peritoneal adhesions due to peritonitis make surgery more difficult and may cause complications. Clarifying the formation mechanism of peritoneal adhesions could help identify methods useful for their prevention. We cultured mesothelial monolayers on plates and microcarriers to simulate the parietal and visceral peritoneum, respectively. We then investigated the effects of lipopolysacchride (LPS) and tumor necrosis factor (TNF) on the homologous adhesion of these mesothelial monolayers. There was no adhesion of mesothelial monolayers in the control medium. When monolayers were cultured with endotoxin (LPS), approximately 90% of the microcarriers adhered to the mesothelial microplate. Adhesions occurred at LPS concentrations of 10 ng/ml and increased linearly in a dose-dependent manner. Kinetic studies revealed that the mesothelial adhesion appeared at 12 hr, and that 90% of the microcarriers were adherent after 24 hr. Open intercellular spaces were observed after a 24-hr treatment with LPS. Scanning electron microscopy revealed that the mesothelial cells adhered to the naked glass. LPS also caused increased permeability of the mesothelial monolayer. TNF did not cause any significant adhesion. Through our experiments we were able to develop an in vitro model of peritoneal adhesion using peritoneal mesothelial cell culture. Endotoxin caused an increase in homologous adhesion of peritoneal mesothelial monolayers, which may correspond to the initial stage of peritoneal adhesion formation in peritonitis.

Dose-Response Relationship, Drug↗
PubMed1995

Molecular evolution of receptors for eicosanoids.

The amino acid sequences of the receptors for various prostaglandins, thromboxane and lipoxin, which belong to the rhodopsin family, were aligned, and a phylogenetic tree was constructed to infer the evolutionary history of the arachidonic acid cascade. The obtained tree suggested that the origin of the cyclooxygenase pathway was different from that of the lipoxygenase pathway. The receptors involved in the cyclooxygenase pathway constructed an independent cluster, but the lipoxin A4 receptor, which is involved in the lipoxygenase pathway, belonged to the cluster of peptide receptors. The primitive form of the cyclooxygenase pathway had been a signal transduction system composed of prostaglandin E2 and its receptor associated with cAMP metabolism.

Amino Acid Sequence↗
PubMed1995

Structural and functional significance of cysteine residues of glutathione-independent prostaglandin D synthase. Identification of Cys65 as an essential thiol.

Glutathione-independent prostaglandin D synthase in rat brain is composed of 189 amino acid residues and catalyzes the isomerization of prostaglandin H2 to prostaglandin D2, an endogenous sleep-promoting substance. This enzyme is the only enzyme among members of the lipocalin superfamily composed of various secretory lipophilic ligand-carrier proteins and is recently identified to be a beta-trace protein, a major constituent of human cerebrospinal fluid. We expressed the active enzyme in Escherichia coli and then systematically substituted all cysteine residues of the delta 1-29 enzyme at positions of 65, 89, and 186 with alanine or serine. The parent and mutant enzymes were purified to apparent homogeneity with a recovery of approximately 30% by chromatography with Sephadex G-50 and S-Sepharose, by which all the enzymes showed identical elution profiles. The purified enzymes, irrespective of the mutation, showed almost the same circular dichroism spectral characteristics as displayed by a highly ordered beta-structure. The recombinant enzymes containing Cys65 showed the activity comparable with that of the enzyme purified from rat brain (approximately 3 mumol/min/mg of protein) in the presence, but not in the absence, of sulfhydryl compounds. However, all of the single, double, and triple mutants without Cys65 lost the enzyme activity. The purified delta 1-29 Ala89,186 enzyme was inactivated reversibly by conjugation with glutathione at Cys65 and irreversibly by the stoichiometric chemical modification with N-ethylmaleimide. These results indicate that Cys65 is an essential thiol of the enzyme and that both the intrinsic and extrinsic sulfhydryl groups are necessary for nonoxidative rearrangement of 9,11-endoperoxide of prostaglandin H2 to produce prostaglandin D2 catalyzed by the enzyme.

Amino Acid Sequence↗
PubMed1995

Expression of blood group-related glycoconjugates in the junctional and other oral epithelia of rodents.

BACKGROUND: The junctional epithelium (JE) attaches the gingiva to the non-vital tooth surface and has other unusual properties which protect the underlying periodontal tissues. The JE differs from other gingival and oral epithelia in its unusual expression of cytokeratins typical of both stratifying and of simple epithelia, a phenotypic pattern possibly related to its specialized functions. METHODS: The patterns of differentiation of rodent gingival and other epithelia were examined using monoclonal antibodies against various glycoconjugates which are expressed on epithelial cell surfaces and provide an alternative marker system for regionally-differing patterns of cell maturation. RESULTS: Markers that are typical of basal cells in other stratifying epithelia were expressed by all cell strata of JE. JE lacked differentiation markers typical of other stratifying oral epithelial but showed suprabasal expression of markers typically expressed by simple epithelia and specialized epithelia, such as taste buds. CONCLUSIONS: The phenotype of rodent JE differs from that of other oral epithelia and the pattern of differentiation assessed by its expression of glycoconjugates parallels that for other phenotypic markers, such as cytokeratins. Differentiation of rodent JE is similar to that of human JE. The functional significance of these patterns of expression is not yet clear but the markers characterizing this unusual epithelium in rodents may be associated with its behavior in periodontal disease and of value to experimental studies of its development.

ABO Blood-Group System↗
PubMed1995

Cloning and sequence analysis of the gene for phosphoenolpyruvate carboxylase from an extreme thermophile, Thermus sp.

The ppc gene, which encodes phosphoenolpyruvate carboxylase (PEPC) of an extreme thermophile, Thermus sp., was cloned and sequenced. The ppc gene had a high G+C content (69.2%). An open reading frame for a 857-amino-acid polypeptide was found in the gene. The calculated molecular mass was 95,632. The amino acid sequence of Thermus PEPC was 31-37% identical and 52-57% similar to those of 17 PEPCs from mesophilic organisms. No Cys residue was found in the polypeptide, demonstrating that this residue is not essential for the catalytic activity of PEPC. The cloned gene was expressed in Escherichia coli and thermostable PEPC was obtained.

Amino Acid Sequence↗
PubMed1995

Cloning, expression and sequence analysis of cDNA for the luciferases from the Japanese fireflies, Pyrocoelia miyako and Hotaria parvula.

Cloning and sequence analysis of cDNA for the luciferases of Pyrocoelia miyako and Hotaria parvula were carried out (GenBank accession numbers L39928 and L39929, respectively). The amino acid sequence, deduced from the nucleotide sequence, showed P. miyako luciferase to consist of 548 amino acid residues with a molecular weight of 60,955, while the luciferase of H. parvula consisted of 548 amino acid residues with a molecular weight of 60,364. Pyrocoelia miyako luciferase showed 82.1% homology with the luciferase of Photinus pyralis and less than 70% homology with other firefly luciferases, whereas H. parvula luciferase showed 98%, 82.5% and 81.2% homology with the luciferases of Luciola mingrelica, Luciola lateralis and Luciola, cruciata respectively. Two regions in the enzymes were found to be highly conserved. The amino acid sequences were used to construct a phylogenetic tree, which showed that the fireflies could be divided into two groups.

Amino Acid Sequence↗
PubMed1995

Molecular evolution of the Ca(2+)-binding photoproteins of the Hydrozoa.

Alignment of the primary structures of the hydrozoan photoproteins, aequorin, mitrocomin, clytin and obelin showed very strong amino acid sequence identities. The Ca(2+)-binding sites of the proteins were found to be highly conserved. The Ca(2+)-binding sites were also homologous to the Ca(2+)-binding sites of other Ca(2+)-binding proteins. However, aequorin, mitrocomin, clytin and obelin differed from other Ca(2+)-binding proteins in that they contained a relatively large number of cysteine, tryptophan, histidine, proline and tyrosine residues, suggesting that these residues may have evolved as part of the light-emitting mechanism. Construction of a phylogenetic tree showed that aequorin, mitrocomin, clytin and obelin form a closely related group of proteins.

Aequorin↗
PubMed1995

Effect of denervation on morphogenesis of the rat fungiform papilla.

In an attempt to elucidate the effects of denervation on development and maintenance of the structure of the fungiform papilla, unilateral neurectomy of the chorda tympani-lingual nerve of rats was performed at day 1 and at weeks 1, 2, 3, 4, 7, and 10 after birth. Specimens were obtained at days 3, 7 and 10, weeks 2, 3, 4, 6, and 8, and months 3 and 4 after neurectomy for examination by light and scanning electron microscopy. At first, the fungiform papillae were atrophic, then progressed to forms resembling filiform papillae. When an immature fungiform papilla was denervated, it eventually changed to papilla identical to normal filiform papillae. The elicited changes differed according to the time of neurectomy; it was found that early neurectomy resulted in a more rapid and marked morphological change of the fungiform papillae. The filiform-like papillae derived from the fungiform ones showed various shapes, sizes, and orientations and were rarely present on the unoperated control side of the lingual dorsum. Sections of the filiform-like papillae revealed that they had no taste buds. These findings suggest: (1) Morphogenesis and structural maintenance of the fungiform papillae require the presence of the chorda tympani and/or lingual nerve. (2) Completion of differentiation and maturation differ in time among fungiform papillae. (3) Fungiform papillae may be transformed filiform papillae induced and maintained by a neurotrophic factor of factors coming from the chorda tympani and/or lingual nerve. (4) Fungiform papillae are rarely innervated contralaterally.

Animals↗
PubMed1994

Membrane-bound Bacillus cytochromes c and their phylogenetic position among bacterial class I cytochromes c.

Gram-positive bacteria lack a periplasmic compartment and contain only membrane-bound cytochromes c. There are at least two types. One is found in subunit II of cytochrome oxidase, and the other is small cytochrome c which is also membrane-bound because of an unprocessed signal sequence or post-translational acylation at the N-terminal end of the protein. These Bacillus cytochromes c are compared with known class I cytochromes c, and a phylogenetic tree has been constructed by the neighbour-joining method.

Amino Acid Sequence↗
PubMed1994

Cloning and expression of the gene for hydroxypyruvate reductase (D-glycerate dehydrogenase from an obligate methylotroph Hyphomicrobium methylovorum GM2.

The gene encoding hydroxypyruvate reductase, catalyzing the asymmetric reduction of hydroxypyruvate to D-glycerate, and its flanking regions were isolated from a methylotrophic bacterium, Hyphomicrobium methylovorum GM2. Nucleotide sequencing of the recombinant plasmids revealed that the hydroxypyruvate-reductase gene codes for the 322-amino-acid protein with calculated molecular mass 35,726 Da. The sequence was confirmed by sequencing the intact enzyme and peptides obtained by digestion of the enzyme with Achromobacter proteinase I. The amino acid sequence of the enzyme showed similarity to members of the D-isomer-specific 2-hydroxyacid dehydrogenase family. The recombinant plasmid, which was constructed by ligation of the cloned gene and an expression vector pKK223-3, was introduced into Escherichia coli HB101. The recombinant enzyme purified from the transformed E. coli cells was indistinguishable from the enzyme isolated from H. methylovorum GM2 by immunological and enzymological analyses.

Alcohol Oxidoreductases↗
PubMed1994

High-level expression of the photorepair gene in Drosophila ovary and its evolutionary implications.

DNA photolyase catalyzes light-dependent repair of cis, syn-cyclobutane dipyrimidines (pyrimidine dimers); its apoenzyme is encoded by the photorepair (phr) gene. The phr cDNA was cloned from D. melanogaster; it has an open reading frame to encode a 61,483-Da protein. The phr cDNA hybridized to band 44C-D of Drosophila polytene chromosome, equivalent to the locus of the phr- gene. Drosophila photolyase is made of an apoenzyme with a molecular weight of 62 kDa. Drosophila photolyase is extraordinarily abundant in the embryo and adult ovary, whereas mRNA of the phr gene is abundant only in the ovary. The action spectrum of Drosophila photolyase for photoreactivation has a maximum at 440 nm. The phr gene of Drosophila has about 60% identical amino acid sites with that of goldfish but only 13-18% with those of microorganisms. Implications of the unique characteristics of the Drosophila phr gene are discussed overviewing the diversified characteristics of phr genes in various organisms that have presumably evolved from a common ancestral gene.

Amino Acid Sequence↗
PubMed1994

Case report: the cleidocervical muscle with speculation as to its origin.

The occurrence of a cleidocervical muscle, which arose from the anterior tubercle of the transverse process of the 6th cervical vertebra and was inserted onto the superior margin of the clavicle, is described. Detailed observations on its innervation, which was derived from the 5th cervical nerve, and its topographic anatomy suggest that the muscle originated from longus colli.

Aged↗
PubMed1993

Molecular evolution of biotin-dependent carboxylases.

Amino-acid sequences of three functional units from various biotin-dependent carboxylases, biotin carboxylase, biotin-carboxyl-carrier protein and carboxyl transferase, were investigated by computer-assisted sequence comparison to obtain information about the structure, function, and molecular evolution of the enzymes. Biotin-dependent carboxylases, except transcarboxylase and oxaloacetate decarboxylase which lack biotin carboxylase, exert their catalytic activities through the three functional units. The three functional units correspond with functional domains or subunits of the enzymes, and the genetic information for the units is encoded in different ways from enzyme to enzyme. It is known that biotin carboxylase is homologous to carbamoyl-phosphate synthetase, and that the biotin-carboxyl-carrier protein is homologous to lipoic-acid-binding domain. The evolutionary relationships between the functional units and their homologues were described. A model for the evolutionary history of the enzymes was proposed by molecular phylogenetic analysis, which shows how a wide variety of domain and/or subunit structures for the enzymes may have been established. A repeated structure was found in biotin-carboxyl-carrier protein, and the secondary structure of the protein was predicted using the observed sequence similarity with a lipoic-acid-binding domain.

Acetyl-CoA Carboxylase↗