ABSTRACT
Background: In regard of chloride channel electrophysiological behavior importance in cellular homeostasis maintenance, some of diseases appearance because of chloride channels impairment, also reports of synchronization between chloride channels impairment and misadjusted pH and that presumably acid or basic pH in cytoplasmic and endoplasmic reticulum luminal spaces are effective on this behavior, current study was performed
Materials and Methods: Research was performed by experimental method. Vesicles from rat liver tissue endoplasmic reticulum were extracted and assessed in 30 samples in 6 groups. Electrophysiological behaviors of channels were measured in control, acidic and basic pH in cis and Trans environments and according of channel conductance and Po this behavior was determined and judged statistically. Data were filtered at 1 kHz and stored at a sampling rate of 10 kHz for offline analysis by PClamp9. Statistical analysis was performed based on Markov noise free single channel analysis
Results: Channel conductance was 72 pS and its current - Voltage relation curve was linear. Channel has Voltage dependent behavior and has grater Po in positive Voltages. Channel conductance in acidic pH remained at 72 pS as of control situation. Channel Po was not changed. In basic pH these findings were also repeated. Also, in cis and Trans spaces these behaviors were sawed
Conclusion: It seems that in pH stream from 6 to 8.5, current channel electrophysiological behavior could be important in endoplasmic reticulum and cellular homeostasis maintenance especially in positive ion such as calcium ion accumulation situation in cytoplasm
ABSTRACT
Defects in endoplasmic reticulum homeostasis are common occurrences in different diseases, such as diabetes, in which the function of endoplasmic reticulum is disrupted. It is now well established that ion channels of endoplasmic reticulum membrane have a critical role in endoplasmic reticulum luminal homeostasis. Our previous studies showed the presence of an ATP-sensitive cationic channel in endoplasmic reticulum. Therefore, in this study, we examined and compared the activities of this channel in control and diabetic rats using single-channel recording techniques. Male Wistar rats were made diabetic for 2 weeks with a single dose injection of streptozotocin [45 mg/kg]. Ion channel incorporation of rough endoplasmic reticulum of diabetic hepatocytes into the bilayer lipid membrane allowed the characterization of K+ channel. Ion channel incorporation of rough endoplasmic reticulum vesicles into the bilayer lipid revealed that the channel current-voltage [I-V] relation with a mean slope conductance of 520 +/- 19 pS was unaffected in diabetes. Interestingly, the channel Po-voltage relation was significantly lower in diabetic rats at voltages above +30 mV. We concluded that the endoplasmic reticulum cationic channel is involved in diabetes. Also, this finding could be considered as a goal for further therapeutic plans
ABSTRACT
Amino acid dehydrogenases [L-amino acid: oxidoreductase deaminating; EC 1.4.1.X] are members of the wider superfamily of oxidoreductases that catalyze the reversible oxidative deamination of an amino acid to its keto acid and ammonia with the concomitant reduction of either NAD[+], NADP[+] or FAD. These enzymes have been received much attention as biocatalysts for use in biosensors or diagnostic kits to screen amino acid metabolism disorders such as phenylketonuria [PKU], maple syrup urine disease [MSUD], homocystinuria [HCY] and hyperprolinemia. This study was aimed to isolation and screening of novel amino acid dehydrogenases from soil bacteria. The enzyme producing bacteria were selected among L-methionine and L-phenylalanine utilizers isolated from soil by thin layer chromatography, activity staining and confirmed by enzyme assay. Bacterial strains were identified by phenotypic and biochemical characteristics. The steady-state kinetic studies of enzymes were also performed. In total of 230 tested strains, four of them were recognized as amino acid dehydrogenase producers that belong to species of Pseudomonas, Citrobacter and Proteus. They exhibited the desired NAD[+]-dependent dehydrogenase activities toward L-isoleucine, L-methionine, L-cysteine, L-serine and L-glutamine in oxidative deamination reaction. The specific activity of L-isoleucine dehydrogenase, L-methionine dehydrogenase and L-glutamine dehydrogenase for oxidative deamination of L-isoleucine, L-methionine and L-glutamine were 1.59, 1.2 and 0.73 U/mg, respectively. The Kcat /Km [s-1.mM -1] values in these strains were as follows: L-isoleucine, 113.6, L-methionine, 62.05 and L-glutamine, 95.83. This is the first report of occurrence a specific isoleucine dehydrogenase, glutamine dehydrogenase and methionine dehydrogenase in bacteria