Purification and some properties of cyclohexylamine oxidase from a Pseudomonas sp.
(9/25)
Cyclohexylamine oxidase was purified 90-fold from cell-free extracts of Pseudomonas sp. capable of assimilating sodium cyclamate. The purified enzyme was homogeneous in disc electrophoresis, and the molecular weight was found to be approximately 80,000 by gel filtration. The enzyme catalyzed the following reaction: cyclohexylamine+O2+H2O leads to cyclohexanone+NH3+H2O2. The enzyme thus can be classified as an amine oxidase; it utilized oxygen as the ultimate electron acceptor. The pH optimum of the reaction was 6.8 and the apparent Km value for cyclohexylamine was 2.5 X 10(-4) M. The enzyme was highly specific for the deamination of alicyclic primary amines such as cyclohexylamine, but was found to be inactive toward ordinary amines used as substrates for amine oxidases. The enzyme solution was yellow in color and showed a typical flavoprotein spectrum; the addition of cyclohexylamine under anaerobic conditions caused reduction of the flavin in the native enzyme. The flavin of the prosthetic group was identified as FAD by thin layer chromatography. The participation of sulfhydryl groups in the enzymic action was also suggested by the observation that the enzyme activity was inhibited in the presence of PCMB and could be recovered by the addition of glutathione. (+info)
Anionic leak currents through the Na+/monocarboxylate cotransporter SMCT1.
(10/25)
Electrical membrane properties of a cell clone derived from human nonpigmented ciliary epithelium.
(15/25)
Intracellular potentials were measured in a SV-40 virus-transformed cell clone derived from human nonpigmented ciliary epithelium using the microelectrode technique. (1) Membrane potential averaged -50.2 mV (+/- 0.6, n = 207). (2) Increasing the extracellular K+ concentration depolarized the membrane voltage. The amplitude of this potential response was reduced in the presence of 1 mM Ba2+. (3) Superfusing the cells with a Ca2+-free solution containing 1 mM EGTA depolarized the intracellular potential and diminished the voltage response upon increasing extracellular K+. (4) Extracellular alkalinization hyperpolarized the membrane potential and increased the voltage amplitude on increasing extracellular K+. (5) Addition of ouabain immediately reduced the intracellular potential. Removing extracellular K+ depolarized membrane voltage, readdition of K+ after K+ depletion transiently hyperpolarized intracellular voltage. Both potential responses were inhibited in the presence of ouabain. (6) Replacing extracellular Cl- by cyclamate resulted in a transient depolarization followed by a hyperpolarization. In the presence of SITS or DIDS (greater than or equal to 0.1 mM) the electrical responses of the cell membrane to Cl- replacement were blocked. We conclude that cultured human nonpigmented ciliary epithelial cells possess an electrogenic Na+/K+-ATPase, a K+ conductance modulated by Ca2+ and pH, and a Cl- conductance sensitive to stilbene derivatives. (+info)
Carcinogen-induced chromosomal breakage decreased by antioxidants.
(16/25)
Blood leukocyte cultures were incubated with antioxidants and the carcinogens sodium cyclamate and 7,12-dimethylbenz(alpha)anthracene in different combinations. There were 17.4% more chromosomal breaks in the group of cells treated with dimethylbenzanthracene only than in the untreated controls. The reductions in chromosomal breaks by the antioxidants were as follows: ascorbic acid, 31.7%; butylated hydroxytoluene, 63.8%; Na(2)SeO(3), 42.0%; and dl-alpha-tocopherol, 63.2%. Multiple chromosomal breaks were distributed equally throughout the experimental groups. Sodium cyclamate had only slightly more chromosomal breaks than the controls (11.6 compared to 10.9%). In the cyclamate groups treated with Na(2)SeO(3), 11.2% of chromosomes were broken. More acrocentric-type chromosomal breaks (21.7%) were seen in the untreated cells than the cells treated with cyclamate (3.4%) or dimethylbenzanthracene alone (4.8%). The carcinogen-treated groups had a higher percentage of meta breaks than the untreated controls. (+info)