ABSTRACT
Acetic acid bacteria Gluconobacter oxydans subsp. industrius RKM V-1280 were immobilized into a synthetic matrix based on polyvinyl alcohol modified with N-vinylpyrrolidone and used as biocatalysts for the development ofbioanodes for microbial fuel cells. The immobilization method did not significantly affect bacterial substrate specificity. Bioanodes based on immobilized bacteria functioned stably for 7 days. The maximum voltage (fuel cell signal) was reached when 100-130 µM of an electron transport mediator, 2,6-dichlorophenolindophenol, was added into the anode compartment. The fuel cell signals reached a maximum at a glucose concentration higher than 6 mM. The power output of the laboratory model of a fuel cell based on the developed bioanode reached 7 mW/m2 with the use of fermentation industry wastes as fuel.
Subject(s)
2,6-Dichloroindophenol/chemistry , Bioelectric Energy Sources , Gluconobacter oxydans/chemistry , Glucose/metabolism , Polymers/chemistry , 2,6-Dichloroindophenol/metabolism , Biocatalysis , Cells, Immobilized , Electrodes , Electron Transport , Fermentation , Gluconobacter oxydans/metabolism , Glucose/chemistry , Industrial Waste , Oxidation-Reduction , Polyvinyl Alcohol/chemistry , Pyrrolidinones/chemistryABSTRACT
Methanol dehydrogenase (MDG) of the facultative methylotrophic phytosymbiont Methylobacterium nodulans has been purified for the first time to an electrophoretically homogeneous state and characterized. The native protein with a molecular mass of 70 kDa consists of large (60 kDa) and small (6 kDa) subunits. The purified protein displayed a specter identical to that of pyrochinolinchinon (PCC)-containing MDGs (pI 8.7, pH optimum in the range 9-10). The enzyme was inactive in the absence of ammonium or methylamine and exhibited a wide substrate specificity with regard to C1-C2 alcohols with the highest affinity to methanol (K(M) = 70 mM), but it did not oxidize benzyl and secondary alcohols. The apparent values of K(M) to primary alcohols increased with the length of the carbonic chain. The enzyme was characterized by a high stability level even in the absence of a substrate. An immobilized enzyme was used for amperometric methanol detection.
Subject(s)
Alcohol Oxidoreductases/isolation & purification , Bacterial Proteins/isolation & purification , Biosensing Techniques , Methanol/analysis , Methylobacterium/enzymology , Protein Subunits/isolation & purification , Alcohol Oxidoreductases/chemistry , Ammonia/chemistry , Bacterial Proteins/chemistry , Crotalaria/microbiology , Electrochemical Techniques , Electrophoresis, Polyacrylamide Gel , Enzyme Stability , Enzymes, Immobilized/chemistry , Enzymes, Immobilized/isolation & purification , Hydrogen-Ion Concentration , Kinetics , Methanol/chemistry , Methylamines/chemistry , Methylobacterium/chemistry , Molecular Weight , Protein Subunits/chemistry , Rhizosphere , Substrate Specificity , SymbiosisABSTRACT
The neuregulin (Nrg) family of growth/differentiation factors is encoded by at least four genes in the mammalian genome: nrg-1, nrg-2, nrg-3 and nrg-4. Nrg-1 and Nrg-2 share the highest homology within the family, and the primary RNA transcripts from their encoding genes are subjected to extensive alternative splicing. Although little is known about the biological function of Nrg-2-4, their structural similarity with Nrg-1 suggests that they could account for some of the activities presently attributed to Nrg-1. Thus, at the neuromuscular junction Nrg-1 has been a favored candidate for the signal that activates selective acetylcholine receptor (AChR) transcription in synaptic myonuclei. However, we have recently shown that like Nrg-1, Nrg-2 can also activate AChR transcription in cultured myotubes and accumulates at the synaptic site. Synapse-specific and Nrg-1-induced AChR transcription require an enhancer sequence, the N-box, which is also mutated in some patients with congenital myasthenia gravis. Here, we show that Nrg-2-induced AChR transcription requires an N-box motif and is regulated by alternative splicing. We also show that unique Nrg-2 isoforms are differentially distributed between spinal cord and skeletal muscle, the tissues that harbor the cellular components of the neuromuscular synapse.
Subject(s)
Alternative Splicing , Nerve Growth Factors/physiology , Receptors, Cholinergic/genetics , Transcription, Genetic , Amino Acid Sequence , Animals , Cloning, Molecular , Consensus Sequence , Humans , Molecular Sequence Data , Muscle, Skeletal/physiology , Nerve Growth Factors/genetics , Peptide Fragments/chemistry , Rats , Schwann Cells/cytology , Schwann Cells/physiology , Sciatic Nerve/physiology , Spinal Cord/physiologyABSTRACT
The study of Ataxia-telangiectasia (A-T) has benefited significantly from mouse models with knockout mutations for the Atm (A-T mutation) locus. While these models have proven useful for in vivo studies, cell cultures from Atm null embryos have been reported to grow poorly and then senesce. In this study, we initiated primary cultures from adult ears and kidneys of Atm homozygous mice and found that these cultures immortalized readily without loss of sensitivity to ionizing radiation and other Atm related cell cycle defects. A mutational analysis for loss of expression of an autosomal locus showed that ionizing radiation had a mutagenic effect. Interestingly, some spontaneous mutants exhibited a mutational pattern that is characteristic of oxidative mutagenesis. This result is consistent with chronic oxidative stress in Atm null cells. In total, the results demonstrate that permanent cell lines can be established from the tissues of adult mice homozygous for Atm and that these cell lines will exhibit expected and novel consequences of this deficiency.
Subject(s)
Ataxia Telangiectasia/genetics , Cell Line, Transformed , Oxidative Stress , Protein Serine-Threonine Kinases/genetics , Radiation, Ionizing , Animals , Ataxia Telangiectasia Mutated Proteins , Cell Cycle/radiation effects , Cell Cycle Proteins , Cell Survival/radiation effects , Chromosome Aberrations , DNA-Binding Proteins , Loss of Heterozygosity/radiation effects , Metaphase/radiation effects , Mice , Mice, Knockout , Mutagenesis , Mutation , Radiation Tolerance , Tumor Suppressor ProteinsABSTRACT
The presence of increased frequencies of blood-derived and solid tumors in ataxia-telangiectasia (A-T) patients, coupled with a role for the ATM (A-T mutation) protein in detecting specific forms of DNA damage, has led to the assumption of a mutator phenotype in A TM-deficient cells. Supporting this assumption are observations of increased rates of chromosomal aberrations and intrachromosomal homologous recombinational events in the cells of A-T patients. We have bred mice with knockout mutations for the selectable Aprt (adenine phosphoribosyltransferase) locus and the Atm locus to examine the frequency of second-step autosomal mutations in Atm-deficient cells. Two solid tissues were examined: (a) the ear, which yields predominately mesenchymal cells; and (b) the kidney, which yields predominately epithelial cells. We report here the lack of a mutator phenotype for inactivating autosomal mutations in solid tissues of the Atm-deficient mice.
Subject(s)
Adenine Phosphoribosyltransferase/genetics , Mutation , Protein Serine-Threonine Kinases , Proteins/genetics , Animals , Ataxia Telangiectasia/genetics , Ataxia Telangiectasia Mutated Proteins , Cell Cycle Proteins , Chromosome Mapping , DNA-Binding Proteins , Ear , Female , Genotype , Heterozygote , Homozygote , Kidney/enzymology , Male , Mice , Mice, Inbred C57BL , Mice, Inbred DBA , Organ Specificity , Proteins/metabolism , Tumor Suppressor ProteinsABSTRACT
To determine the types of mutations induced by oxidative damage, a kidney cell line with a heterozygous deficiency for the autosomal Aprt (adenine phosphoribosyltransferase) gene was tested for its mutagenic response to hydrogen peroxide. Aprt-deficient cells were selected and scored for loss of heterozygosity (LOH) for 11 microsatellite loci on mouse chromosome 8. On the basis of the LOH analysis, spontaneous mutants (n = 38) were distributed into four classes: apparent point mutation, mitotic recombination, chromosome loss, and large interstitial deletion. However, 9 of 20 (45%) hydrogen peroxide-induced mutants exhibited a novel class of mutations characterized by "discontinuous LOH" for one or more of the microsatellite loci. Interestingly, mutations resembling discontinuous LOH are commonly observed in a wide variety of human cancers. Our data suggest that discontinuous LOH is a signature mutational pattern for oxidative damage and further suggest that such genetic damage is widespread in cancer.
Subject(s)
Adenine Phosphoribosyltransferase/genetics , Loss of Heterozygosity , Mutation , Neoplasms/genetics , Animals , Cells, Cultured , Chromosomes, Human, Pair 8 , DNA Damage/genetics , Female , Humans , Hydrogen Peroxide/pharmacology , Kidney/cytology , Male , Mice , Mice, Inbred C57BL , Mutation/drug effects , Oxidative Stress/geneticsABSTRACT
Ultrastructure of the serotonin-producing (EC) duodenum cells of a hibernant in euthermia (in summer and winter) and hypothermia has been studied. It has been found that EC cells of an active summer and spontaneously arousal animals belong to active population. This cells have protein-synthetic activity. The winter cells are characterized by the deficiency of the synthetic processes and the secretory granules disintegration. The results support the hypothesis that ES cells are an important element in the regulation of the hibernation.