Search PubMed⌕ Search

Biomedical subjects

K Hamaguchi

Publications and source records attributed to K Hamaguchi.

At least 199 records · Page 11Linked to original sources
PubMed1986

Conformation and stability of the constant fragment of the immunoglobulin light chain containing an intramolecular mercury bridge.

The constant fragment of the immunoglobulin light chain in which the intramolecular disulfide bond is reduced (reduced CL fragment) assumes a conformation very similar to that of the intact CL fragment and contains two sulfhydryl groups buried in the interior of the molecule [Goto, Y., & Hamaguchi, K. (1979) J. Biochem. (Tokyo) 86, 1433-1441]. In order to understand the role of the disulfide bond, a derivative in which the disulfide bond is replaced by an S-Hg-S bond was prepared and its conformation and stability were studied. The derivative was prepared by reacting the reduced CL fragment with mercuric chloride. Kinetic studies showed that the reaction is rate-limited by the unfolding process of the reduced CL fragment. The mercury derivative was as compact as the intact CL or reduced CL fragment, and a tryptophyl residue was found to be buried near the S-Hg-S bond in the interior of the protein molecule. Judging from the circular dichroic spectrum, however, the beta-structure characteristic of the immunoglobulin fold was disturbed. The stability of the derivative to guanidine hydrochloride was lower than that of the intact CL fragment, but the unfolding transition was reversible and cooperative. Decreased stability of the mercury derivative is due to its folded conformation being distorted by introduction of the S-Hg-S bond.

Bence Jones Protein↗
PubMed1986

Reduction of the buried intrachain disulfide bond of the constant fragment of the immunoglobulin light chain: global unfolding under physiological conditions.

The constant (CL) fragment of the immunoglobulin light chain contains only one intrachain disulfide bond buried in the interior of the molecule. The kinetics of reduction with dithiothreitol of the disulfide bond were studied at various concentrations of guanidine hydrochloride at pH 8.0 and 25 degrees C. It was found that the disulfide bond is reduced even in the absence of guanidine hydrochloride. The results of the reduction kinetics were compared with those of the unfolding and refolding kinetics of the CL fragment previously reported [Goto, Y., & Hamaguchi, K. (1982) J. Mol. Biol. 156, 891-910]. It was shown that the reduction of the disulfide bond proceeds through a species with a conformation very similar to that of the fully unfolded one and that the CL fragment undergoes global unfolding transition even in water.

Disulfides↗
PubMed1986

Immunoreactive somatostatin and calcitonin in pulmonary neuroendocrine tumor.

A well-differentiated neuroendocrine carcinoma of the lung that secreted immunoreactive somatostatin (IR-SRIF) and IR-calcitonin (CT) in a 72-year-old women is described. The plasma concentrations of IR-SRIF (57.5 pg/ml) and IR-CT (340 pg/ml) before operation were significantly higher than the respective normal ranges. After resection of the tumor, the plasma CT level (105 pg/ml) decreased to within the normal range, and the SRIF level (32.7 pg/ml) also decreased, but was still abnormally higher, which suggested the presence of an unidentified remnant of the tumor. Abnormal accumulation of technetium 99m (99mTc) in the lumbar vertebrae was found 6 months after the operation, which indicated a metastatic tumor. The tissue concentrations of IR-SRIF and IR-CT were 103 and 94 ng/g wet weight, respectively, and SRIF-IR tumor cells and CT-IR tumor cells were demonstrated immunohistochemically. On gel-filtration chromatography of the tumor tissue, two peaks of SRIF immunoreactivity were eluted in the positions of synthetic SRIF-28 and SRIF-14, respectively. Conversion of SRIF-28 to SRIF-14 was suggested from results on changes in the two IR-SRIF components during incubation with a crude enzyme preparation extracted from the tumor tissue.

Aged↗
PubMed1986

Isolation of glucagon-secreting cell lines by cloning insulinoma cells.

Six glucagon-secreting cell lines designated as In-R1-G1, -G3, -G7, -G9, -G10, and -G11 were isolated from insulinoma cells (In-111-R1) by single cell cloning. A small amount of insulin was also detectable in the incubation medium when hormone secretion was stimulated by the addition of arginine or theophylline. These cell lines grew as monolayers and the population doubling times varied from 16.8 to 28.8 h. Karyologically these clones were aneuploid and the modes of chromosome numbers were 61 to 70. Electron microscopic examination of one of these clones showed that these cells contained moderately developed Golgi apparatus and a few secretory granules, which more or less resembled alpha-cell granules. By gel filtration study of the incubation medium, glucagon and glucagonlike material were eluted. The molecular weight of the latter was approximately 9000, which suggested the concomitant secretion of proglucagon into the medium. The levels of secreted glucagon in basal state were 0.3 to 3.0 ng/10(6) cells/2 h. Glucagon secretion was markedly enhanced in the presence of amino acids. Glucagon secretion increased slightly in the presence of high concentration of glucose in Hanks' balanced salt solution; however it was not affected by the varying concentrations of glucose when the cells were incubated in complete media with amino acids. Glucagon secretion was also stimulated by the addition of theophylline. These clonal cell lines seem to provide a useful tool for investigating the mechanism of glucagon secretion.

Adenoma, Islet Cell↗
PubMed1986

Cellular hypersensitivity to nervous antigens in Guillain-Barré syndrome.

Cell-mediated immune responses to various nervous antigens were examined in 12 cases of Guillain-Barré syndrome (GBS), 24 cases of noninflammatory peripheral neuropathy (NIPN), and 18 cases of degenerative disorders of central nervous system (CNSDD), using the lymphocyte-transformation technique. Cellular hypersensitivity to bovine P2 protein (P2) and a synthetic peptide, SP66-78, corresponding to the residues 66-78 of P2, was detected in about two-thirds of GBS cases, especially in the active or improving stages, but not in NIPN and CNSDD. The lymphocytes sensitized to these nervous antigens might play an important role in the pathogenesis of GBS.

Antigens↗
PubMed1986

Unfolding by temperature and guanidine hydrochloride of chicken pancreatic polypeptide.

Thermal unfolding of chicken pancreatic polypeptide at two different concentrations was studied at various pH values. The thermal stability was higher at higher protein concentrations. The transition temperatures at two different protein concentrations changed with pH in parallel and decreased by about 30 degrees C on lowering pH from 5 to 2. The results on the thermal unfolding were analyzed by assuming that the dimerization constant is independent of pH, that the thermal unfolding occurs only after the pancreatic polypeptide dimers dissociated into the monomers, and that one ionizable group participates in the acid unfolding of the monomer. The free energy change for the unfolding of the pancreatic polypeptide monomer was estimated to be 1.4 kcal/mol. The unfolding of pancreatic polypeptide by guanidine hydrochloride at pH 6.0 and 25 degrees C was also studied. The stability to guanidine hydrochloride was higher at higher protein concentrations.

Animals↗
PubMed1986

Unfolding and refolding of the constant fragment of the immunoglobulin light chain containing an intramolecular mercury bridge.

The conformation of the constant fragment of the immunoglobulin light chain in which the intrachain disulfide bond is replaced by the bond S-Hg-S (CL-Hg fragment), was as compact as that of the intact CL fragment, but its stability to guanidine hydrochloride was lower than that of the intact CL fragment [Goto, Y. & Hamaguchi, K. (1986) Biochemistry in press]. The kinetics of reversible unfolding and refolding of the CL-Hg fragment by guanidine hydrochloride were studied and compared with those for the intact CL and reduced CL fragments [Goto, Y. & Hamaguchi, K. (1982) J. Mol. Biol. 156, 891-910, 911-926]. The unfolding kinetics were explained on the basis of a three-species mechanism, U1----U2----F, where U1 and U2 are respectively slow-folding and fast-folding species of unfolded protein, and F is folded protein. However, an additional isomerization, though its contribution to the overall reaction process is small, had to be taken into account to explain the refolding kinetics. The kinetic properties of interconversion between U1 and U2 were similar to those for the intact CL and reduced CL fragments. This suggested that the same prolyl residue is involved in the isomerization reactions in the unfolded states of the intact CL, reduced CL, and CL-Hg fragments. The rate constant for the unfolding process, F to U2, was about 20 times greater than those for the intact CL and reduced CL fragments, while the rate constant for the refolding process, U2 to F, lay between the values for the intact CL and the reduced CL fragment. The free energy profiles of unfolding and refolding of the intact CL, reduced CL, and CL-Hg fragments were compared.

Bridged-Ring Compounds↗
PubMed1986

Combined effects of glucose and oxygen concentrations upon the ultrastructure of cultured islet cells and insulin accumulation in culture media.

Rat pancreatic islets cultured for 6 days at 100 or 500 mg/dl glucose and 20 or 7% O2 were examined electron-microscopically, and insulin accumulation in the culture media was assayed immunologically. In the islets cultured at 500 mg/dl glucose and 20% O2, B cells exhibited hypertrophy of granular endoplasmic reticulum and Golgi apparatus, an abundance of free ribosomes, degranulation and the margination of secretory granules. In islets cultured at 500 mg/dl glucose and 7% O2, B cells exhibited dilatation of endoplasmic reticulum cisternae and dominance of Golgi vesicles in addition to the above-mentioned changes. These changes, together with the correlated data on insulin accumulation, are discussed with special reference to the effects of glucose and oxygen upon the synthesis and release of insulin in B cells.

Animals↗
PubMed1985

pH-induced unfolding of the constant fragment of the immunoglobulin light chain: effect of reduction of the intrachain disulfide bond.

Circular dichroism and proton nuclear magnetic resonance measurements were made to clarify how reduction and reduction plus alkylation of the intrachain disulfide bond affect the conformation and stability of the constant (CL) fragment of the immunoglobulin light chain. The pH titration behavior of the two histidine residues in the intact CL, reduced CL, and reduced and alkylated CL fragments were followed by proton nuclear magnetic resonance spectroscopy. It was shown that reduction of the intrachain disulfide bond does not affect the solution conformation of the immunoglobulin fold, whereas reduction and alkylation results in extensive unfolding of the protein molecule. These results are consistent with the previous results obtained by Goto and Hamaguchi [1979) J. Biochem. 86, 1433-1441) using circular dichroism, fluorescence, and titration of SH groups. The pH-induced unfolding of the reduced CL fragment was compared with that of the intact CL fragment. The stability of the intact CL fragment to acid was much greater than that of the reduced CL fragment. The equilibria of the unfolding by acid could be explained by assuming that the same ionizable groups participate in the unfolding for the reduced CL fragment as for the intact CL fragment, and that the stability of the intact CL fragment is 100 times greater than that of the reduced CL fragment. In the alkaline pH region, the reduced CL fragment was unfolded above pH 8.5, while the intact CL fragment was not unfolded until pH 11.6. The kinetics of the unfolding by alkali of the reduced CL fragment was also studied. The unfolding by alkali of the reduced CL fragment could be explained by assuming that the reduced CL molecule can no longer adopt the folded conformation when either or both SH groups ionize.

Circular Dichroism↗
PubMed1984

Cysteinyl residues of Escherichia coli recA protein.

The Escherichia coli recA protein has three cysteinyl residues at positions 90, 116, and 129. All of them are reactive with 5,5'-dithiobis(2-nitrobenzoic acid) (DTNB). In the presence of ATP or ADP, only one cysteinyl residue reacts with DTNB. The residue was also reactive with N-[7-(dimethylamino)-4-methylcoumarinyl]maleimide (DACM) in the presence of ATP. The results on an analysis of the DACM-modified protein cleaved at the nonmodified cysteinyl residues after cyanation with 2-nitro-5-(thiocyano)benzoic acid show that two cysteinyl residues protected in the presence of ATP or ADP are identified as Cys-90 and Cys-129. When the ionic strength is higher than 1, one cysteinyl residue does not react with DTNB. This residue is Cys-90 or Cys-129, because one of the two cysteinyl residues, which are not modified with DACM in the presence of ATP, does not react with DTNB at high ionic strength. The binding of single-stranded DNA to the recA protein does not change the reactivity of the cysteinyl residues with DTNB.

Amino Acids↗
PubMed1984

2',3'-Cyclic nucleotide 3'-phosphodiesterase activity in the cerebrospinal fluid of patients with demyelinating diseases.

We aimed to study the level of CNPase activity in the cerebrospinal fluid of patients with demyelinating diseases and other neurological diseases, particularly multiple sclerosis, with reference to CSF myelin basic protein content. CNPase activity was measured paper chromatographically using radioactive 2',3'-cAMP as a substrate. Myelin basic protein content was measured with a radioimmunoassay. The mean level of CNPase activity was significantly higher for multiple sclerosis than for nonneurological controls. Dividing the disease phases of multiple sclerosis into the three periods, the CNPase activity was found to be significantly elevated in the worsening period and reduced in the improving period and the inactive period. The level of CNPase activity in the cerebrospinal fluid of multiple sclerosis coincided with the clinical activity of the disease. The level of CNPase activity correlated well (r = 0.84) with the level of myelin basic protein content in cerebrospinal fluid. The ratio for CNPase activity and myelin basic protein content in cerebrospinal fluid was almost the same as that in human central nerve myelin. We concluded that CNPase activity in the cerebrospinal fluid from neurological patients is an indicator of destruction of myelin in the central nervous system, and the measurement of CNPase activity in the cerebrospinal fluid of multiple sclerosis could be useful in the clinical management.

2',3'-Cyclic Nucleotide 3'-Phosphodiesterase↗