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Biomedical subjects

M Yazawa

Publications and source records attributed to M Yazawa.

At least 19 recordsLinked to original sources

Calcium dependent conformational changes of surfactant protein A (SP-A) and its collagenase resistant fragment with or without dithiothreitol.

Calcium-dependent conformational changes of surfactant protein A (SP-A) and the collagenase resistant fragment (CRF) of SP-A were studied by measuring fluorescence spectra. The emission peaks of both SP-A and CRF in the absence of Ca2+ appeared at 343 nm when they were excited at 280 nm. In the presence of Ca2+, the peaks appeared at 340 nm and were accompanied by an increase in the fluorescence intensity. The magnitude of the fluorescence intensity change induced by Ca2+ was amplified by the addition of dithiothreitol (DTT) in both SP-A and CRF. The Ca2+ binding of CRF was measured by a flow dialysis method with 45CaCl2 in the Ca2+ concentration range where the Ca(2+)-induced fluorescence changes occurred. The maximum binding number of Ca2+ to CRF was about 2 mol per mol of CRF, and the value was independent of the presence of DTT.

Calcium

Amino acid sequence of calmodulin from Euglena gracilis.

The complete amino acid sequence of calmodulin from Euglena gracilis was determined by isolation and sequence analyses of peptides derived from calmodulin by digestion with trypsin and Staphylococcus aureus V8 protease. Euglena calmodulin consists of 148 amino acid residues; it lacks tryptophan and cysteine and contains one tyrosine, three histidine and two NE-trimethyllysine residues/molecule of the protein. Its N-terminus was blocked with an acetyl group and C-terminal lysine was trimethylated. Euglena calmodulin is the first calmodulin so far examined in which the C-terminal lysine is trimethylated. The comparison of amino acid sequences between Euglena and human brain calmodulins indicated 17 amino acid substitutions in Euglena calmodulin.

Amino Acid Sequence

Binding of calcium by calmodulin: influence of the calmodulin binding domain of the plasma membrane calcium pump.

The interaction between calmodulin and synthetic peptides corresponding to the calmodulin binding domain of the plasma membrane Ca2+ pump has been studied by measuring Ca2+ binding to calmodulin. The largest peptide (C28W) corresponding to the complete 28 amino acid calmodulin binding domain enhanced the Ca2+ affinity of calmodulin by more than 100 times, implying that the binding of Ca2+ increased the affinity of calmodulin for the peptide by more than 10(8) times. Deletion of the 8 C-terminal residues from peptide C28W did not decrease the affinity of Ca2+ for the high-affinity sites of calmodulin, but it decreased that for the low-affinity sites. A larger deletion (13 residues) decreased the affinity of Ca2+ for the high-affinity sites as well. The data suggest that the middle portion of peptide C28W interacts with the C-terminal half of calmodulin. Addition of the peptides to a mixture of tryptic fragments corresponding to the N- and C-terminal halves of calmodulin produced a biphasic Ca2+ binding curve, and the effect of peptides was different from that on calmodulin. The result shows that one molecule of peptide C28W binds both calmodulin fragments. Interaction of the two domains of calmodulin through the central helix is necessary for the high-affinity binding of four Ca2+ molecules.

Amino Acid Sequence

[A case of surgery for annuloaortic ectasia and aortic regurgitation complicated by ulcerative colitis and aortitis syndrome].

We performed an operation for AAE and AR complicated by ulcerative colitis and aortitis syndrome. The patient was a 20-year-old male who had been treated for ulcerative colitis in our hospital since 1983, when he was 18 years old. In 1985, he was admitted to our hospital for treatment and evaluation of left heart failure. He was diagnosed as having AAE and AR due to aortitis syndrome, and steroid therapy was started. He developed heart failure, and surgery was indicated. At operation, before clamping the aorta we made a composite graft. The ascending aorta and aortic valve were replaced by the composite graft, and button-shaped coronary ostia were sutured directly into the graft. His postoperative course was uneventful and he was discharged. He is now maintained on steroid therapy for his aortitis syndrome.

Adult

[Autonomic nerve disorders in generalized amyloidosis].

Various autonomic disturbances are usually seen in systemic amyloidosis with polyneuropathy, especially in familial amyloid polyneuropathy (FAP). In this paper we summarized the clinicopathological features of these autonomic symptoms. Orthostatic hypotension and bowel dysfunctions are two major autonomic manifestations of FAP, and at autopsy severe deposition of amyloid is observed in an extensive area of peripheral autonomic nervous system including sympathetic ganglia. Remarkable depletion of the extrinsic nerves with relative preservation of the intrinsic nerves is a characteristic finding in the gastrointestinal tract of the patients with FAP, and this abnormal innervation may produce the peculiar bowel disorders. Oral administration of L-threo-3,4-dihydroxyphenylserine, a precursor of noradrenaline, is effective for the treatment of these autonomic symptoms of FAP patients.

Adult

[Spontaneous intercostal lung hernia].

A 70-year-old man admitted our clinic because of a painful bulge in the left chest wall that had appeared following a bout of coughing. CT confirmed the presence of lung herniation at the left eight interspace. At surgery a 3 x 16 cm anterolateral defect in the site was confirmed. A patch of Gore-Tex (expanded polytetrafluoroethylene) was fashioned and sewn into the defect with interrupted 3-0 Ethibond sutures. Spontaneous intercostal lung hernia is a rare lesion of thoracic wall and have not been reported in Japan. We describe a case with some reviewing.

Aged

Noradrenergic nerve fibers of the rectal mucosa in autonomic disorders: comparison of histochemical study with clinical severity and changes in plasma noradrenaline induced by standing.

A histochemical study was carried out on the rectal mucosae biopsied from 20 patients with autonomic dysfunctions and 13 controls using a catecholamine fluorescent staining method, and the rectal noradrenergic nerve fiber lesions were compared with the severity of autonomic symptoms and disturbance of plasma noradrenaline increase in response to standing in 17 patients. In 9 patients with type I familial amyloid polyneuropathy and 1 with acute pandysautonomia, the number of fluorescent nerve fibers was greatly reduced, and the degree of depletion correlated well with the other 2 parameters showing the severity of autonomic dysfunction. In contrast, rectal noradrenergic nerve fibers were normally preserved in 10 patients with multiple system atrophy, although they suffered from severe autonomic symptoms with poor noradrenaline response to the postural loading. It is concluded that noradrenergic nerve fiber lesions in the biopsied rectal mucosa may represent the systemic involvement of sympathetic post-ganglionic nerves.

Adult

A site-directed mutagenesis study of yeast calmodulin.

A site-directed mutagenesis study was carried out in order to understand the regulatory mechanism of calmodulin. We started from the yeast (Saccharomyces cerevisiae) calmodulin gene since it has many differences in amino acid sequence and inferior functional properties compared with the vertebrate calmodulin. Recombinant yeast calmodulins were generated in Escherichia coli transformed by constructed expression plasmids. Three recombinant calmodulins were obtained. The first two were YCM61G, in which the Ca2(+)-binding site 2 (the four Ca2(+)-binding EF-hand structures in calmodulin were numbered from the N-terminus) was converted to the same as that in vertebrate calmodulin, and YCM delta 132-148, in which the C-terminal half sequence of site 4 was deleted. These two recombinant calmodulins had the same maximum Ca2+ binding (3 mol/mol) as yeast calmodulin, which indicates that site 4 of yeast calmodulin was the one losing Ca2+ binding capacity. YCM delta 132-148 could not activate target enzymes, whereas its Ca2+ binding profile was similar to those of yeast calmodulin and YCM61G. Therefore, the structure in site 4 which cannot bind Ca2+ is indispensable for the regulatory function of yeast calmodulin. The complete regulatory function of vertebrate calmodulin can be attained by the combination of 4 Ca2+ binding structures. The negative charge cluster in the central alpha-helix region is suggested to stabilize the active conformation of calmodulin, since the third yeast calmodulin mutant, YCM83E, which had the negative charge cluster, increased the maximum activation of myosin light chain kinase.

Amino Acid Sequence

Monoclonal antibodies toward scallop (Patinopecten yessoensis) testis and wheat germ calmodulins.

A monoclonal antibody (IM7) toward scallop testis calmodulin and another one (PBE2) toward wheat germ calmodulin were produced. Ca2+ was required for IM7 to react with scallop calmodulin. IM7 reacted with the C-terminal region (Asp78-Lys148) of the calmodulin. As observed on competitive ELISA, IM7 reacted with chicken calmodulin, but not with Euglena gracilis or wheat calmodulin, troponin C, myosin light chains, or parvalbumin. It is assumed that the cluster of Thr143, Thr146, and Ser147 in the C-terminal region acts as the antigenic site. IM7 (and Fab of IM7) inhibited the activities of myosin light chain kinase and cAMP-phosphodiesterase. PBE2 reacted with wheat germ calmodulin irrespective of the presence or absence of Ca2+, the antigenic site being in the N-terminal region (Ala1-Met37). It reacted with wheat and spinach calmodulins, but not with scallop, chicken, or Euglena calmodulin, troponin C, myosin light chains, or parvalbumin. PBE2 had no effect on the activities of myosin light chain kinase and cAMP-phosphodiesterase.

3',5'-Cyclic-AMP Phosphodiesterases

Mastoparan binding induces Ca(2+)-transfer between two globular domains of calmodulin: a 1H NMR study.

The interaction between calmodulin and mastoparan at various concentrations of calcium ions was studied by 1H NMR. It was found that at lower mastoparan concentrations 1 mol of mastoparan binds to both the C-terminal-half and N-terminal-half regions of calcium-saturated calmodulin. The mastoparan affinity is much greater for the C-terminal-half region than for the N-terminal-half region. At higher mastoparan concentrations, a further 1 mol of mastoparan binds to the N-terminal-region of calcium saturated calmodulin. The results can be interpreted in terms of the assumption that the N-terminal-half region of calmodulin with mastoparan has a higher calcium ion affinity than the C-terminal-half region without mastoparan. It is suggested that calcium ions transfer from the C-terminal-half region of calmodulin without mastoparan to the N-terminal-half region of calmodulin with mastoparan. This calcium ion transfer is discussed from the viewpoint of enzyme activation by calmodulin.

Amino Acid Sequence

1H NMR study on amide proton exchange of calmodulin-mastoparan complex.

Amide proton exchange rates of Ca2(+)-saturated calmodulin and Ca2(+)-saturated calmodulin-mastoparan complex were studied by 1H NMR spectroscopy. Exchange rates of Gly25, Gly61, Gly98, Gly134, Ile27, Ile100, and Asn137 were determined for Ca2(+)-saturated calmodulin and for Ca2(+)-saturated calmodulin-mastoparan complex, and were found to be less than 10(-4)s-1. All these residues of which the amide proton resonances appear at lower fields were considered to form hydrogen bonds, based on the results of X-ray analysis. Exchange rates of Ile27 and Asn137 became an order of magnitude smaller when mastoparan bound to Ca2(+)-saturated calmodulin, while those of the four glycines and Ile100 did not change appreciably. The reduction in accessibility of Asn137 to water cased by mastoparan binding suggests that a part of the mastoparan binding site is probably located in or near the hydrophobic cluster of the C-terminal-half domain. The reduction in accessibility of Ile27 also suggests that another part of the mastoparan binding site is located in or near the hydrophobic cleft of the N-terminal-half domain.

Amino Acids

[Mediastinal parathyroid cyst].

A 66-year-old woman, having no complaint, admitted our clinic, because of a mass in the right superior mediastinum detected three years ago on chest X-ray. The operation was performed through the right thoracotomy in December of 1989. The thin-walled cyst located adjacent to the trachea between superior vena cava and back bones, occupied from beneath the innominate artery to the right main bronchus. The tumor (7 x 5 x 5 cm) weighing 80 grams had a smooth surface and contained watery fluid. Histological examination showed a cyst lined a monolayer of cuboidal epithelium. The cyst wall consisted of parathyroid tissue. Mediastinal parathyroid cyst is very rare. Since the first report of DeQuervain, 15 cases have been reported. These reports are reviewed.

Aged

[Broncholithiasis: clinical experience of three operated cases].

Three patients with broncholithiasis had an operation in our hospital. Patient 1 had no history of tuberculosis. She had the pulmonary stones removed bronchoscopically twice. The stones were composed of inflammatory granulomas. Patient 2 had a history of pulmonary tuberculosis. Endoscopical removal of the pulmonary stone was not successful. After 6 days, she coughed up a large amount of blood, and the operation was performed. Patient 3 coughed up blood for 10 years. The amount of blood gradually increased and the operation was performed. All three cases had calcification of the hilar lymph nodes. Broncholithiasis appears to be related not only to pulmonary tuberculosis but also to other chronic inflammatory diseases of the lung.

Adult

Inter-domain interaction and the structural flexibility of calmodulin in the connecting region of the terminal two domains.

The calcium-dependent difference absorption spectrum of scallop calmodulin was measured in the presence of mastoparan. The difference spectrum at 286 nm (delta A286) showed biphasic response to Ca2+ concentration. The first change represents the conformational change around Tyr-138 and the second change may respond to an interaction between N- and C-domain of calmodulin which became apparent in the associated state with mastoparan. Calmodulin-mastoparan complex was eluted from a gel filtration column after free calmodulin in the presence of Ca2+, which indicates a more compact structure of calmodulin-mastoparan complex than of free calmodulin. The biphasic response of delta A286 was also observed with free calmodulin when the ionic strength was as low as 0.02 M NaCl. In the absence of NaCl, the Ca2+ dependence of delta A288 was monophasic, assuming identical affinity of Ca2+ to both domains. Increase in the sensitivity of calmodulin to trypsin was observed with decrease in ionic strength. These results suggest an ionic-strength-dependent decrease in ordered structure of the connecting region. Calmodulin may change shape depending upon the ionic strength by bending at the connecting region. We assumed from the observations that calmodulin in solution may fluctuate between the two extreme shapes of the bent and the dumbbell structure. Target proteins may select and fix the specific bent structure for their activation.

Animals

Histone H1 kinase specific to the SPKK motif.

A protein kinase phosphorylating sea urchin spermatogenous histones, H1 and H2B, was found in sea urchin egg homogenate and purified. The kinase is activated by cAMP and is composed of two different types of subunits with molecular masses 41 and 46 kDa. The kinase phosphorylates a peptide, Ser-Pro-Arg-Lys-Ser-Pro-Arg-Lys, which is a double repeat of the DNA-binding SPKK motif [Suzuki M., (1989) EMBO J. 8, 797-804]. We name this kinase SPkinase because it exclusively phosphorylates H1 and H2B, the only histones containing SPKK motifs. Phosphorylation of H1 by SPkinase decreases the DNA-binding ability of H1. This paper is the first to report purification of a kinase which affects the DNA-binding ability of a gene regulatory protein.

Amino Acid Sequence

[A case of B-cell non-Hodgkin's lymphoma with prominent splenomegaly and high serum titer of cold agglutinin].

The case of 37-year-old male with non-Hodgkin's lymphoma with prominent splenomegaly is presented. The spleen that was removed for analysis had a weight of 2690 grams and a section inspection revealed multiple, whitish, small nodules that were disseminated throughout the entire spleen. It was microscopically demonstrated to be a B-Cell lymphoma of diffuse, medium-sized cells and an IgM.kappa Type. This case also was complicated by a high serum titer of cold agglutinin that was decreased by chemotherapy.

Adult

113Cd-NMR evidence for cooperative interaction between amino- and carboxyl-terminal domains of calmodulin.

113Cd-NMR experiments were performed to characterize the nature of Cd2+ binding to calmodulin in the presence of a tetradecapeptide mastoparan or a 26-residue peptide M13 (calmodulin-binding region of skeletal muscle myosin light-chain kinase). The results indicate that binding of these peptides to calmodulin induces a positive cooperativity between Ca2+ binding to C- and N-terminal domains. The results imply that the activation of myosin light-chain kinase caused by the increase in Ca2+ concentration occurs as a result of cooperative interactions not only between two Ca2+ binding sites in each domain but also between the two domains. The interdomain interaction manifests itself only in the presence of such peptides.

Animals