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Biomedical subjects

T Akino

Publications and source records attributed to T Akino.

At least 109 records · Page 6Linked to original sources
PubMed1990

Farnesylated gamma-subunit of photoreceptor G protein indispensable for GTP-binding.

Transducin, composed of subunits T alpha, T beta and T gamma, is a member of a heterotrimeric G-protein family, and transduces the light signal in visual cells. We have recently found that bovine T beta gamma can be separated into two components. T beta gamma-1 and T beta gamma-2, each of which has its own gamma-subunit, T gamma-1 and T gamma-2, respectively. T beta gamma-2 enhances the binding of GTP to T alpha in the presence of metarhodopsin II by about 30-fold compared with T beta gamma-1. Here we show that a farnesyl moiety is attached to a sulphur atom of the C-terminal cysteine of T gamma-2 (active form), a part of which is additionally methyl-esterified at the alpha-carboxyl group. In T gamma-1 (inactive form), however, such modifications are missing. Thus, the farnesyl moiety attached to the gamma-subunit is indispensable for the GTP-binding activity of transducin. This suggests that a similar modification may occur in the gamma-subunits of other heterotrimeric G proteins involved in biological signal transduction processes.

Amino Acid Sequence↗
PubMed1990

D-amino acids in mouse tissues are not of microbial origin.

Neutral free D-amino acid contents in the serum, kidney, liver, brain, small intestine and urine in germ-free mice and those in specific pathogen-free mice were compared. No significant difference was found. This strongly suggests that the free D-amino acids which were shown to be present in mice in our previous work did not originate from the enteric microbial flora.

Amino Acids↗
PubMed1990

Evidence for coupling of different receptors for gonadotropin-releasing hormone to phospholipases C and A2 in cultured rat luteal cells.

Effects of [D-Ala6,Des-Gly10]gonadotropin-releasing hormone (GnRH), ethylamide (GnRHa), and prostaglandin F2 alpha (PGF2 alpha) on inositol phosphate (IPs) formation and arachidonic acid (AA) release were studied in rat luteal cells of primary culture. In the cells obtained from one-day-old corpora lutea, PGF2 alpha (100 nM) and GnRHa (100 nM) significantly increased the IPs formation and the AA release. Antagonists of GnRH added solely or with GnRHa did not stimulate the IPs formation but did stimulate the AA release. In the cells obtained from 5-day-old corpora lutea, GnRHa failed to stimulate the IPs formation but significantly stimulated the AA release. The stimulation of both IPs formation and AA release by PGF2 alpha was consistently found in cells of two different luteal ages. These results suggest that GnRH receptor independently couples to both phospholipases C and A2 through different classes of GnRH receptors.

Animals↗
PubMed1990

A specific beta gamma-subunit of transducin stimulates ADP-ribosylation of the alpha-subunit by pertussis toxin.

Transducin (T alpha and T beta gamma) is a GTP-binding protein involved in the visual transduction process in a rod outer segment. We have previously demonstrated that T beta gamma is a mixture composed of two components, T beta gamma-1 and T beta gamma-2, with distinctive gamma-subunits, T gamma-1 and T gamma-2, respectively (Fukada et al., 1989, J. Biol. Chem., 264, 5937-5943). To investigate the interaction between T alpha and the two components of T beta gamma, the effect of either T beta gamma-1 or T beta gamma-2 on the ADP-ribosylation of T alpha catalyzed by pertussis toxin (IAP) was examined. T beta gamma-2 stimulated the ADP-ribosylation of T alpha by IAP, while T beta gamma-1 displayed almost no enhancement of the ADP-ribosylation. Addition of T beta gamma-1 to the mixture of T alpha and T beta gamma-2 had no effect on the ADP-ribosylation of T alpha. These results indicate that T alpha and T beta gamma-2 form a complex that serves as a substrate of IAP in the ADP-ribosylation reaction, while T beta gamma-1 has a little affinity for T alpha. It was suggested that T gamma-2 is an essential subunit for T beta to interact with T alpha.

Adenosine Diphosphate Ribose↗
PubMed1990

Binding of GTP to transducin is not inhibited by arrestin and phosphorylated rhodopsin.

In the presence of a photobleaching intermediate of unphosphorylated or phosphorylated rhodopsin (Rh*), the binding of GppNHp to transducin was measured with or without arrestin for elucidation of the shut-off mechanism of the visual transduction process in bovine rod outer segments. The ability of Rh* to catalyze the formation of the transducin-GppNHp complex in the absence of arrestin was independent of the degree of phosphorylation of Rh*. Furthermore, the catalyzing ability of the phosphorylated Rh* was not reduced by the addition of arrestin. These observations indicate that the interaction between phosphorylated Rh* and transducin was not inhibited by arrestin. Thus, the hypothesis was not supported that the PDE shut-off process is a simple competition between transducin and arrestin for binding to phosphorylated Rh*.

Animals↗
PubMed1990

Molecular species profiles of acidic phospholipids in lung fractions of adult and perinatal rabbits.

The molecular species of phosphatidylglycerol (PG) and phosphatidylinositol (PI) in pulmonary surfactant and membrane fractions of adult and perinatal rabbit lungs were analyzed by high-performance liquid chromatography of the dinitrobenzoyl derivatives of the diacylglycerols (DGs), derived from the two acidic phospholipids. The PG in both surfactant and membrane fractions of adult lungs consisted mainly of the 16:0/16:0 species, followed in order by 16:0/18:1 and 16:0/18:2 species. In contrast, the prominent molecular species of PI in the membrane fractions were 18:0/20:4 and 16:0/18:1, while surfactant PI consisted mainly of 16:0/18:1 and 16:0/18:2, containing only 3% of 16:0/16:0. In the perinatal rabbit lungs, a compositional change between surfactant PG and PI was found, i.e., an increase in PG and a decrease in PI. The molecular species compositions of PG and PI in the perinatal lungs were generally the same as those observed in the adult lungs. It should, therefore, be noted that the species profiles of surfactant PG and PI, particularly in the content of 16:0/16:0 and 18:0/20:4, are not similar, but distinctly different from each other in both adult and developing lungs. Therefore, the present results strongly suggest two possibilities; (1) both acidic phospholipids which appear in pulmonary surfactant may originate from different pools of CDP-DGs having different molecular species; and/or (2) surfactant PG and PI may be synthesized by individual enzymes having different substrate specificities for different CDP-DGs in alveolar type II cells.

Animals↗
PubMed1990

Functional heterogeneity of beta gamma-subunit of frog transducin.

1. Transducin subunits (T alpha and T beta gamma) were purified from freshly dissected frog (Rana catesbeiana) retinas. It was found that purified T beta gamma is composed of three components which can be separated from each other by an anion exchange column chromatography under nondenaturing conditions. 2. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analyses of these three components demonstrated that each contains T beta (mol. wt 35,000) and T gamma (mol. wt approximately 8000). 3. Only one of the three components retained an ability to enhance the binding of GppNHp to T alpha in the presence of a photobleaching intermediate of rhodopsin, while the others showed very low abilities to enhance the binding. 4. These observations, together with the similar findings on bovine T beta gamma, strongly suggest that the functional heterogeneity of T beta gamma is conserved in vertebrate photoreceptor cells.

Animals↗
PubMed1990

Pulmonary corpora amylacea contain surfactant apoprotein.

The case of a 53-year-old female with interstitial pneumonitis is described with special regard to biochemical characterization of pulmonary corpora amylacea which were found in the lung specimen obtained by bronchial biopsy from the patient. The main protein component in bronchoalveolar lavage (BAL) fluid of the patient was albumin, but proteins in the precipitate fraction of BAL fluid, where the corpora amylacea were recovered, predominantly consisted of 36 kD protein which was stained with the monoclonal antibody PE 10 to human pulmonary surfactant apoprotein by immunoblot. Histologically the pulmonary corpora amylacea were stained with eosin and PAS. The particles were stained immunohistochemically by immunoperoxidase reaction using PE 10, but not by antibodies to human albumin. The pulmonary surfactant apoprotein seems, therefore, to be not simply adsorbed in the particles, but to be contained in them. Thus, the surfactant apoprotein may, at least in this case, be involved in the formation of pulmonary corpora amylacea.

Apoproteins↗
PubMed1989

Characterization of phospholipids accumulated in pulmonary-surfactant compartments of rats intratracheally exposed to silica.

Phospholipids in the lung fractions, i.e. alveolar free cells, extracellular pulmonary surfactant, intracellular pulmonary surfactant (lamellar bodies) and microsomal fractions, of rats were examined 28 days after intratracheal injection of silica (40 mg/kg). Significant accumulations of phospholipids were observed in the extracellular- and intracellular-surfactant fractions of rats exposed to silica. The prominent phospholipid accumulated was phosphatidylcholine (PC), consisting mainly of the dipalmitoyl species. However, a compositional change in acidic phospholipids of surfactant fractions was produced by the silica treatment. The percentage of phosphatidylglycerol (PG) was significantly decreased; in contrast, that of phosphatidylinositol (PI) was increased. Thus the ratio PG/PI in the surfactant fractions was markedly decreased in response to silica. This compositional change in both acidic phospholipids occurred even in the early stages, i.e. before appreciable accumulations of alveolar phospholipids were noticed. The molecular-species profiles of both acidic phospholipids in the surfactant fractions were distinctly different from each other. The dipalmitoyl species accounted for more than 30% of PG and less than 6% of PI, respectively. These species patterns of PG and PI were similar in control and silica-treated rats. These findings suggest two possibilities that (1) PG and PI destined for pulmonary surfactant are synthesized from each specific CDP-diacylglycerol (DG) pool having different molecular species in the lung, or (2) individual enzymes responsible for synthesis of surfactant PG and PI have substrate specificities for molecular species of CDP-DG, thereby producing PG and PI having different molecular species in surfactant compartments.

Animals↗
PubMed1989

Alterations of acidic phospholipids in bronchoalveolar lavage fluids of patients with pulmonary alveolar proteinosis.

We analyzed phospholipids, especially the acidic phospholipids, i.e. phosphatidylglycerol (PG) and phosphatidylinositol (PI), of bronchoalveolar lavage (BAL) fluids from patients with pulmonary alveolar proteinosis. A significant decrease in PG and a concomitant increase in PI were found in the phospholipid of the patient's BAL fluids compared to that of normal subjects. Thus, the PG to PI ratio was significantly decreased in pulmonary alveolar proteinosis. These changes in the acidic phospholipids in BAL fluids of patients with pulmonary alveolar proteinosis appear to indicate that a switch-over in the synthesis of both acidic phospholipids from a same precursor CDP-diacylglycerol (CDP-DG) in alveolar Type II cells occurs in a such diseased state. However, since the molecular species profiles of PG and PI in the patient's BAL fluids were distinctly different from each other, the PG and PI present in the alveoli of the patients appeared to be derived from different pools of CDP-DG in alveolar Type II cells.

Bronchoalveolar Lavage Fluid↗
PubMed1989

The presence of free D-amino acids in mouse tissues.

The presence of free D-amino acids in mouse kidney, liver, brain, heart, lung, thymus and serum has been shown with an enzymic microdetermination method. The D-amino acid levels were higher in the extracts of kidney and liver than in those from other organs.

Amino Acids↗
PubMed1989

Beta gamma-subunit of bovine transducin composed of two components with distinctive gamma-subunits.

During the process of transduction of a photon signal in vertebrate rod outer segments, transducin, a guanine nucleotide binding protein, mediates between a photobleaching intermediate of rhodopsin and a cGMP-phosphodiesterase. We report here that the beta gamma-subunit of bovine transducin (T beta gamma) characterized so far consists of two components (T beta gamma-1 and T beta gamma-2), which can be separated by anion exchange chromatography under nondenaturing conditions. Both components consisted of two polypeptides of Mr 36,000 (T beta) and about 8,000 (T gamma) in sodium dodecyl sulfate polyacrylamide (13%) gel electrophoresis. On a further analysis by 8 M urea/sodium dodecyl sulfate-polyacrylamide gel electrophoresis, T gamma subunits of T beta gamma-1 and T beta gamma-2 showed Mr values of 8,000 (T gamma-1) and 6,000 (T gamma-2), respectively. Amino acid compositions of both T gamma-1 and T gamma-2 roughly corresponded with that of T gamma previously reported and were quite different from that of gamma-subunit of cGMP-phosphodiesterase. Western blot analysis of freshly isolated rod outer segments by an antiserum raised against a mixture of T beta gamma-1 and T beta gamma-2 revealed the presence of both components in the membranes of a starting material. This observation excludes the possibility that one of the components might be produced artificially in the course of the purification. In the presence of a photobleaching intermediate of either unphosphorylated or phosphorylated rhodopsin, the binding of guanosine 5'-(beta, gamma-imido)triphosphate (GppNHp) to the alpha-subunit of transducin (T alpha) was remarkably enhanced with increasing concentrations of purified T beta gamma-2. On the contrary, T beta gamma-1 retained little ability, if any, to enhance the GppNHp binding to T alpha; the ability of T beta gamma-1 was at least 30 times lower than that of T beta gamma-2. Such a low activity of T beta gamma-1 was attributed to inability for coupling of T alpha with a photobleaching intermediate of rhodopsin. These results indicate that T gamma-2 is essential for the GTP binding of transducin. The role of T gamma-1 in vertebrate photoreceptor cells was discussed.

Adenosine Triphosphate↗
PubMed1989

Involvement of D-amino acid oxidase in elimination of free D-amino acids in mice.

The physiological role of D-amino acid oxidase was investigated by using mutant ddY/DAO- mice lacking the enzyme. Free D-amino acid concentrations in the mutant mice were significantly higher than those of control ddY/DAO+ mice in kidney, liver, lung, heart, brain, erythrocytes, serum and urine. The results suggest that the enzyme is involved in the catabolism of free D-amino acids in the body, and that free D-amino acids are also excreted into urine.

Amino Acids↗
PubMed1989

Surfactant proteins in the diagnosis of fetal lung maturity. II. The 35 kd protein and phospholipids in complicated pregnancy.

The major surfactant protein with a molecular weight of 35 kd and also saturated phosphatidylcholine and phosphatidylglycerol were analyzed in specimens of amniotic fluid; 68 were from cases of maternal diabetes, 41 from preeclampsia or maternal hypertension, 26 from premature rupture of the fetal membranes, and 45 from normal pregnancies. The relationship between the individual surfactant components was studied after covariance adjustment for the length of gestation. In severe early-onset preeclampsia, the 35 kd surfactant protein/saturated phosphatidylcholine ratio was significantly higher than in the other pregnancies. In diabetic pregnancies (classes B to D without preeclampsia), the phosphatidylglycerol/saturated phosphatidylcholine ratio was lower than in the other pregnancies. Isolated surfactant complex showed similar abnormalities. In severe early-onset preeclampsia and insulin-dependent diabetes without vascular disease, the phosphatidylglycerol/saturated phosphatidylcholine ratio correlated negatively with fetal growth. In four samples of amniotic fluids from cases of severe early-onset preeclampsia, the 35 kd protein falsely predicted lung maturity. All had abnormally high 35 kd protein/saturated phosphatidylcholine ratios (greater than 2 SD of controls). According to the present results, the 35 kd protein may give a false mature test result in severe preeclampsia.

Amniotic Fluid↗
PubMed1989

Two Bacillus beta-mannanases having different COOH termini are produced in Escherichia coli carrying pMAH5.

The nucleotide sequence was determined for the alkalophilic Bacillus sp. strain AM-001 beta-mannanase gene which produced two beta-mannanases (A and B) in Escherichia coli transformants. The putative beta-mannanase gene was 1,539 base pairs long and encoded a mature beta-mannanase protein of 487 amino acids and a signal peptide of 26 amino acids. The COOH-terminal amino acid of beta-mannanase A is an arginine residue located at amino acid 513 of the deduced amino acid sequence, and that of beta-mannanase B is a valine residue located at amino acid 365. Deletion derivatives having 1,098 base pairs from the ATG start codon maintained the beta-mannanase activity of the encoded polypeptide. However, clones harboring DNA fragments (1,051 base pairs) shorter than the gene which encoded beta-mannanase B (1,095 base pairs) did not exhibit the beta-mannanase activity. The simultaneous production of both beta-mannanases A and B in an E. coli transformant was demonstrated by the maxicell procedure.

Amino Acid Sequence↗
PubMed1989

Improved immunoassay for the determination of surfactant protein A (SP-A) in human amniotic fluid.

A simple, improved immunoassay for the determination of human surfactant protein A (SP-A) in human amniotic fluid was developed. The immunoreaction with a monoclonal antibody PC6-immobilized plastic bead, peroxidase-labeled monoclonal antibody PE10 and amniotic fluid sample diluting in the buffer containing 0.6% sodium dodecyl sulfate/2% Triton X-100, was carried out simultaneously at 45 degrees C for 30 min in test tubes. After washing the bead with 2% skimmilk in phosphate buffered saline containing 1% Triton X-100, the peroxidase reaction was developed by adding the substrate reagent and the absorbance was measured. The amniotic fluid obtained at full term was used as a standard instead of purified SP-A, because of the stability of the antigenicity. This immunoassay method was used to measure SP-A in 69 samples of amniotic fluid from 22 to 41 weeks of gestation. The result indicated that the SP-A values obtained by the present immunoassay can be used for predicting the fetal lung maturity. This simplified monoclonal immunoassay completed the measurement within 1 hr, and so it could be used routinely in clinical laboratory.

Amniotic Fluid↗
PubMed1988

Immunohistochemical localization of pulmonary surfactant apoproteins in various lung tumors. Special reference to nonmucus producing lung adenocarcinomas.

Eighty-nine primary lung carcinomas and 23 metastatic lung tumors were immunohistochemically studied for the expression of pulmonary surfactant apoproteins, by using monoclonal (PE-10) and polyclonal antibodies. Surfactant apoprotein was demonstrated in the cytoplasm and/or nuclear inclusion bodies of only primary lung adenocarcinomas (36 of 75 cases), not in any other histologic type of primary lung carcinoma or in metastatic lung tumors. In primary lung adenocarcinoma, although typical type II pneumocyte type adenocarcinoma was not included in the current series, the majority of surfactant apoprotein-positive single cell type tumors were of the Clara cell type, with a single bronchial surface epithelial cell type, according to the light microscopic subclassification of adenocarcinoma cells. The Clara cell type adenocarcinomas could at times be distinguished only with difficulty from adenocarcinoma of type II pneumocyte type. Normal and hyperplastic type II pneumocytes were of course positive for surfactant apoprotein in the cytoplasm. However, none of the positive cells could definitely be identified as Clara cells in non-neoplastic lungs. The findings obtained in this study indicate that surfactant apoprotein is a good marker to distinguish adenocarcinoma of the lung from other histologic types of lung cancer and from neoplasms metastatic to the lung, and that type II pneumocytes and Clara cells, non-neoplastic and neoplastic, are morphologically and functionally closely related and might belong to the same cell lineage.

Adenocarcinoma↗