Search PubMed⌕ Search

Biomedical subjects

M Fujioka

Publications and source records attributed to M Fujioka.

At least 163 records · Page 9Linked to original sources
PubMed1992

Mammalian small molecule methyltransferases: their structural and functional features.

Structural and functional features of mammalian S-adenosyl-methionine-dependent small molecule methyltransferases are reviewed. The methyltransferases have similar protomer molecular weights in the range of 25,000-35,000. Two common sequence motifs are found in all enzymes of known sequence. Whereas the kinetic mechanisms may be different, the methyltransferases in the free form bind S-adenosylmethionine. Most, if not all, of mammalian small molecule methyltransferases appear to have vicinal thiols in a catalytically important area of the enzyme.

Amino Acid Sequence↗
PubMed1992

Rat gro/melanoma growth-stimulating activity. Assessment of the structure responsible for chemotactic activity by use of its fragments prepared by proteolysis and chemical synthesis.

Rat gro/melanoma growth-stimulating activity is a dimer composed of two identical subunits. Each subunit consists of 72 amino-acid residues and contains two disulfide bridges. In order to obtain information on the structure responsible for chemotactic activity, various fragments of gro were prepared and tested for their ability to induce chemotaxis. None of the fragments corresponding to residues 1-6, 1-21, 12-31, 36-50 or 52-72 was active as a chemoattractant. Reduced and carboxymethylated gro as well as the tryptic peptide consisting of three peptides, residues 9-21, 28-45 were and 49-61, linked by two disulfide bonds Cys-9-Cys-35 and Cys-11-Cys-51, were inactive. Also, these, peptides did not inhibit the chemotactic activity of gro. Rat gro lacking the N-terminal 6 residues had a reduced activity and the one lacking the C-terminal Lys was as active as intact gro. Therefore, an almost entire portion of the molecule including disulfide cross-links is required for chemotactic activity.

Amino Acid Sequence↗
PubMed1992

Atelosteogenesis type 3: the first patient in Japan and a survivor for more than 1 year.

We report the first patient of atelosteogenesis type 3 (AO3) in Japan. The patient had multiple craniofacial abnormalities at birth, including ocular hypertelorism, a flat nasal bridge, micrognathia and a cleft palate. There was rhizomelic shortness of the limbs and a club-foot. The infant had short broad thumbs in the hands similar to those observed in the feet. There were no chromosomal abnormalities. Radiological examination demonstrated striking hypoplasia of the humerus with proximal rounding and distal tapering giving a 'drumstick' appearance, 'S'-shape configuration of the cervical spine, scoliosis and coronal cleft in the thoracolumbar vertebral bodies. The infant experienced recurrent apnea and persistent severe tracheomalacia, which necessitated tracheostomy at 5 months of age. Despite his multiple skeletal deformities and respiratory problems, this patient survived more than 1 year with motoneuronal developmental delay.

Abnormalities, Multiple↗
PubMed1992

Multiple endocrine neoplasia type 2A.

A 40-year-old woman was admitted with complaints of headache, palpitation and diaphoresis. She had undergone right hemithyroidectomy 12 years previously. Histological reexamination of the operative specimen revealed a medullary thyroid carcinoma. Abdominal ultrasonography, CT scan and angiography showed bilateral adrenal tumors. Serum catecholamine levels in both adrenal veins were high. Based on these data, bilateral adrenalectomy was performed. Histological examination confirmed the diagnosis of pheochromocytomas. After operation, serum calcitonin and urinary noradrenaline levels were still high. Further examination by 131I-metaiodobenzylguanidine (MIBG) scintigraphy is planned.

Adrenal Gland Neoplasms↗
PubMed1992

[Pneumonectomy in a case of chronic hemodialysis].

A 60-year-old male was referred to our hospital due to a well-delineated tumor (4 x 4 x 3 cm) in the left hilar region demonstrated by chest X-ray examination. Bronchofiberscopic biopsy showed evidence of well moderately differentiated epidermoid carcinoma. Since the tumor infiltrated to a part of the pericardium, left pneumonectomy and partial resection of pericardium were performed. The disease was p-T2, N1, M0, and state II. Anemia was corrected with Erythropoietin, and hemodialysis was performed for 3 consecutive days before operation. Hemodialysis 3 times per week was initiated from 2 days after operation. Hyperkalemia was successfully treated by GI therapy (continuous intravenous infusion of glucose+insulin). His postoperative course was good without bleeding and infection. He was discharged 37 days after operation and has been observed on an outpatient basis. With an increased and aging patients on chronic hemodialysis, the operations for various malignant tumors have been increasing. However, there are few reports on operation for lung cancer and no report on pneumonectomy in such patients in Japan. We performed pneumonectomy for lung cancer detected in a patient on chronic hemodialysis for 4 years and obtained good results.

Carcinoma, Squamous Cell↗
PubMed1991

Nutritional regulation and tissue-specific expression of the serine dehydratase gene in rat.

The mechanism of dietary regulation and tissue-specific expression of the serine dehydratase gene in rat has been studied. The hepatic serine dehydratase activity and its mRNA showed a parallel increase with increasing protein content in the diet. However, when rats that had been maintained on a high protein diet were fed a protein-free diet, the mRNA level rapidly decreased to 0.5 in 3 h, whereas the enzyme activity gradually fell to a low level over a period of 5 days. With animals maintained on a high protein diet or on a protein-free diet, we examined the sites hypersensitive to DNase I in the 5'-flanking region of serine dehydratase gene in the liver chromatins. A series of DNase I-hypersensitive sites were located within 10.5 kilobase pairs upstream of the transcription start site. The DNA regions at -3050 and -3180 (region II) and -3600 to -3850 (region III) were more susceptible to the nuclease in the expressing than in the nonexpressing liver. A reverse situation obtained at -100 (region I). Kidney contained serine dehydratase mRNA at a level of 5% of liver as determined by Northern blotting. The kidney chromatin was found to be susceptible to DNase I only at region I. No conspicuous DNase I-hypersensitive sites were observed in the relevant regions of chromatins from brain and lung, in which serine dehydratase mRNA was scarcely transcribed. These results suggest that nutritional control and tissue-specific expression of the serine dehydratase gene is closely associated with the alteration of DNase I hypersensitivity at specific sites of the 5'-flanking region of the gene.

Animals↗
PubMed1991

Reversible inactivation of recombinant rat liver guanidinoacetate methyltransferase by glutathione disulfide.

Recombinant rat liver guanidinoacetate methyltransferase is inactivated by glutathione disulfide (GSSG) following pseudo-first-order kinetics. A second-order rate constant of 20.8 M-1 min-1 is obtained at pH 7.5 and 30 degrees C. The inactivation is fully reversed by glutathione (GSH) in a pseudo-first-order fashion with a second-order rate constant of 11.1 M-1 min-1. The rate of inactivation is not affected by S-adenosylmethionine or guanidinoacetate, but complete protection against inactivation is observed in the presence of sinefungin plus guanidinoacetate. At equilibrium in the buffers containing various concentrations of GSH and GSSG, the enzyme shows activities that are dependent on the ratio but not on the total concentration of GSH and GSSG. A hyperbolic relationship is obtained between enzyme activity and [GSH]/[GSSG] ratio. The inactivation by GSSG is associated with the disappearance of approximately 1 mol of sulfhydryl group per mole of enzyme. These results indicate that inactivation of guanidinoacetate methyltransferase by GSSG is the consequence of the formation of a mixed disulfide between a protein thiol and glutathione. The equilibrium constant for the redox reaction, E-SH + GSSG in equilibrium with E-SSG + GSH, obtained from the equilibrium data (1.69) is in good agreement with the value determined as the ratio of second-order rate constants for reactivation and inactivation (1.87). The cysteine residue engaged in the mixed disulfide with glutathione is identified as Cys-15 by peptide analysis after consecutive treatment of the GSSG-inactivated enzyme with N-ethylmaleimide, 2-mercaptoethanol, and [14C]iodoacetate. The GSSG-inactivated enzyme binds S-adenosyl-methionine but not guanidinoacetate in the presence and absence of sinefungin. Native guanidinoacetate methyltransferase binds guanidinoacetate in the presence of sinefungin. The low overall redox equilibrium constant of 1.7-1.9 found for the reaction between guanidinoacetate methyltransferase and GSSG suggests that the activity of the enzyme is not amenable to modulation by the change in intracellular [GSH]/[GSSG] ratio.

Adenosine↗
PubMed1991

Rat liver guanidinoacetate methyltransferase. Proximity of cysteine residues at positions 15, 90 and 219 as revealed by site-directed mutagenesis and chemical modification.

Cys-90 of rat liver guanidinoacetate methyltransferase is a very reactive residue, and chemical modification of this residue results in a large decrease in activity [Fujioka, Konishi & Takata (1988) Biochemistry 27, 7658-7664]. To understand better the role of Cys-90 in catalysis, this residue was replaced with alanine by oligonucleotide-directed mutagenesis. The mutant is active and has kinetic constants similar to those of wild-type, indicating that Cys-90 is not involved in catalysis and substrate binding. The u.v.-absorption, fluorescence and c.d. spectra are also unchanged. Reaction of the mutant with an equimolar amount of 5,5'-dithiobis-(2-nitrobenzoic acid) or 2-nitro-5-thiocyanobenzoic acid results in an almost quantitative disulphide cross-linking between Cys-15 and Cys-21). The same treatment effects disulphide bond formation between Cys-15 and Cys-90 in wild type [Fujioka, Konishi & Takata (1988) Biochemistry 27, 7658-7664]. Since the mutant and wild-type enzymes appear to have similar secondary and tertiary structures, these results suggest that Cys-15, Cys-90 and Cys-219 of the methyltransferase occur spatially close together. The mutant cross-linked between Cys-15 and Cys-219 and the wild-type cross-linked between Cys-15 and Cys-90 show very similar spectroscopic properties. Although treatment of the mutant and wild-type enzymes with equimolar concentrations of 5,5'dithiobis-(2-nitrobenzoic acid) causes a large loss of enzyme activity in each case, kinetic analyses with the modified enzymes suggest that cross-linking of Cys-15 with Cys-90 or Cys-219 does not abolish activity and does not result in a large change in the Michaelis constants. Incubation of the mutant enzyme with excess 2-nitro-5-thiocyanobenzoic acid leads to modification of Cys-207 in addition to Cys-15 and Cys-219. Retention of considerable enzyme activity in the modified enzyme indicates that Cys-207 is also not an essential residue.

Amino Acid Sequence↗
PubMed1991

Pig lens glutathione S-transferase belongs to class Pi enzyme.

Class Pi glutathione S-transferase was purified to homogeneity from pig lens cytosol. This enzyme was composed of two identical 22 kDa subunits and had isoelectric point of 8.5 from the results of SDS gel electrophoresis, gel filtration, amino acid sequence analysis and isoelectric focusing. Amino acid sequence of N-terminal 15 residues was almost identical to class Pi enzymes from human, rat and mouse. Antibody against the pig enzyme crossreacted to human glutathione S-transferase-pi and anti-rat glutathione S-transferase-P antibody crossreacted to pig enzyme.

Amino Acid Sequence↗
PubMed1991

Requirement for the beta,gamma-pyrophosphate bond of ATP in a stage between transcription initiation and elongation by Escherichia coli RNA polymerase.

A linear fragment of DNA was fixed to acrylamide or agarose beads by its ends. When a fragment containing the lambda PR promoter is immobilized and transcribed, the RNA products are unchanged from those obtained on the unfixed DNA. Transcription from the immobilized fragment can be interrupted by diluting the reaction mixture into a large volume of the same buffer. Brief centrifugation allows isolation of the transcription complex with the immobilized DNA. If interruption occurs during elongation, the elongation can be resumed upon a second addition of substrates. If ATP is replaced by a beta, gamma-unhydrolyzable analogue in the second addition, the elongated products are similar to those obtained when the substrate contain ATP. When ATP is replaced by the analogue at the initiation step, however, the yield of elongated products is decreased to less than one-sixth and that of short abortive products is increased. Thus the ATP analogues are good substrates once elongation has been established in the presence of ATP, but not good enough to get past a stage just after initiation in the absence of ATP. We conclude that the beta, gamma-pyrophosphate bond of ATP is important for preparation of efficient elongation.

Adenosine Triphosphate↗
PubMed1991

Recombinant rat guanidinoacetate methyltransferase: structure and function of the NH2-terminal region as deduced by limited proteolysis.

Recombinant rat liver guanidinoacetate methyltransferase, a monomeric protein with Mr 26,000, is inactivated upon incubation with low concentrations of trypsin. Examination of the reaction products by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and high-performance liquid chromatography followed by amino acid analysis and sequencing of isolated peptides reveals that the inactivation is due to the cleavage of the NH2-terminal segment after Arg20. The cleaved peptide is not tightly associated with the rest of the protein. The rate of inactivation is not affected by the presence of either S-adenosylmethionine (AdoMet) or guanidinoacetate, but a substantial retardation of inactivation is observed when both substrates are present. The cleavage at Arg20 is also slowed by cross-linking Cys15 and Cys90 by a disulfide bond. An equilibrium binding study shows that guanidinoacetate methyltransferase in the free form binds AdoMet but not guanidinoacetate. The trypsin-modified enzyme, despite having no catalytic activity, can weakly bind AdoMet and guanidinoacetate in the presence of AdoMet. Chymotrypsin rapidly hydrolyzes the peptide bond after Trp19, and elastase cleaves the bond after Ala24, leading in both cases to loss of activity. The results obtained in this study suggest that the portion of the methyltransferase around residues 19-24 is highly exposed to the solvent and flexible. The results also indicate that the NH2-terminal region is not directly involved in substrate binding but plays a role in catalysis.

Amino Acid Sequence↗
PubMed1991

[Leiomyosarcoma of the esophagus associated with pulmonary edema by the compression of the left atrium].

Leiomyosarcoma of the esophagus is a rare neoplasm. We reported a very rare case of esophageal leiomyosarcoma associated with pulmonary edema by the compression of the left atrium. A 67-year-old man was admitted with a 3-month history of chest pain, exertional dyspnea and dysphagia. Chest X-ray computed tomography showed posterior mediastinal tumor. Esophageogastroscopy and fiberoptic bronchofiberscopy showed no direct invasion of the tumor. By echocardiography, the left atrium was found to be compressed by the tumor and pulmonary hypertension (58/25 mmHg) and increased pulmonary wedge pressure (25 mmHg) was present. Open biopsy specimen demonstrated elongated cells suggestive of sarcoma. At autopsy, the tumor was confirmed to be leiomyosarcoma and to be originated from the lower esophagus.

Aged↗
PubMed1991

[A case of mitral stenosis developing infective endocarditis 18 months after PTMC].

We presented here a case of mitral stenosis developing infective endocarditis 18 months after PTMC. A 33-year-old male was admitted to our hospital for the evaluation of mitral stenosis with signs of congestive heart failure. The use of PTMC was adequately indicated based on the data of previous cardiac catheterization and was successfully performed. The post-operative clinical course was good leaving only slight mitral regurgitation demonstrated by post-valvuloplasty LVG. About 18 months after PTMC, the patient developed a high fever and erythema with tenderness on his leg, so he was admitted again. Blood culture was positive for streptococcus viridans. Moreover, echocardiogram revealed valvular vegetation on the anterior mitral leaflet, which is the characteristic finding of infective endocarditis. Antibiotics were so effective that his clinical course was relatively good with minor cerebral infarction during chemotherapy. Little is known about the prognosis and late complication of PTMC. However, our reported case may suggest that prophylactic chemotherapy might be recommended at least in some cases showing mitral regurgitation after PTMC.

Adult↗