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Öğe Carbonic anhydrase activity from the gilthead sea bream (Sparus aurata) liver: The toxicological effects of heavy metals(2013) Kaya E.D.; Söyüt H.; Beydemir T.Many studies have shown that metal ions may lead to oxidative stress in biological systems. Accordingly, DNA damage, protein modification, enzyme inhibition and activation, lipid peroxidation and many other effects may occur in living organisms. Many different formations of metal ions may enter human cells along with water, air, and various foods, and humans are negatively affected by these conditions, either directly or indirectly. These effects may cause irreversible damage to human metabolism. In this study, the toxicological effects of heavy metals on carbonic anhydrase enzyme activity from the gilthead sea bream liver were investigated. The carbonic anhydrase enzyme was purified via affinity chromatography and had a specific activity of 6775.5EUmg-1. The kinetics and characteristic properties, such as optimum pH, stable pH, optimum temperature, activation energy (Ea), activation enthalpy (?H), Q10, Km, and Vmax, were determined for the purified enzyme SDS-polyacrylamide gel electrophoresis showed a single band and molecular weight of the subunit was approximately 25kDa. Cd(II), Cu(II), Ni(II) and Ag(I) inhibited the enzyme activity in vitro. The type of inhibition and Ki values for these metals were calculated from Lineweaver-Burk plots as 17.74mM, 36.20mM, 12.85mM and 0.025mM for Cd(II), Cu(II), Ni(II) and Ag(I), respectively. All the metals were noncompetitive inhibitors. © 2013 The Authors.Öğe Changes in carbonic anhydrase activity and gene expression of Hsp70 in rainbow trout (Oncorhynchus mykiss) muscle after exposure to some metals(2012) Söyüt H.; Beydemir S.; Ceyhun S.B.; Erdo?an O.; Kaya E.D.The effects of some heavy metals Co(II), Cu(II), Zn(II), and Ag(I) on carbonic anhydrase (CA) activity from rainbow trout (RT) muscle were investigated. Moreover, the effects of these metals on the expression of heat shock protein 70 (Hsp70) gene in muscle tissue were examined by real-time quantitative PCR (RT-PCR) in muscle tissue aft er exposure to the metals at the end of 6, 12, 24, and 48 h. CA was purified with a specific activity of 2300 EU mg-1, a yield of 19%, and 1080-fold. The molecular weights (Mw) of subunit and native enzyme were approximately 30 and 31 kDa, respectively. Optimum pH, stable pH, optimum temperature, activation energy (E?), activation enthalpy (?H), and Q10 (the difference in activity of enzyme caused by the increment of 10 oC) value were determined. Apparent Michaelis constant (Km), maximum reaction rate (Vmax), and turnover rate of the enzyme (Kcat) values were 1.29 mM, 0.17 ?mol min-1, and 28.8 s-1, respectively. The catalytic efficiency (Kcat/Km) was 22.3. The heavy metals decreased in vitro CA activity. Inhibition mechanisms of the metal ions were noncompetitive, except for the Co(II) ion, which was competitive. The expression of the Hsp70 gene was increased in the presence of the metal ions. The expression level at the end of 48 h was the highest for all of the metals. Consequently, the in vitro inhibition rank order was determined as Co(II) > Zn(II) > Cu(II) > Ag(I). Interestingly, Ag(I) was the most effective metal ion on Hsp70 gene expression. © TÜBİTAK.Öğe Impact of antibacterial drugs on human serum paraoxonase-1 (hPON1) activity: An in vitro study(Asian Pacific Tropical Biomedicine Press, 2014) Söyüt H.; Kaya E.D.; Beydemir S.Objective: To investigate the in vitro effects of the antibacterial drugs, meropenem trihydrate, piperacillin sodium, and cefoperazone sodium, on the activity of human serum paraoxonase (hPON1). Methods: hPON1 was purified from human serum using simple chromatographic methods, including DEAE-Sephadex anion exchange and Sephadex G-200 gel filtration chromatography. Results: The three antibacterial drugs decreased in vitro hPON1 activity. Inhibition mechanisms meropenem trihydrate was noncompetitive while piperacillin sodium and cefoperazone sodium were competitive. Conclusions: Our results showed that antibacterial drugs significantly inhibit hPON1 activity, both in vitro, with rank order meropenem trihydrate piperacillin sodium cefoperazone sodium in vitro. © 2014 by the Asian Pacific Journal of Tropical Biomedicine.Öğe The toxicological impacts of some heavy metals on carbonic anhydrase from gilthead sea bream (Sparus aurata) gills(Elsevier, 2015) Kaya E.D.; Söyüt H.; Beydemir Ş.It is known that heavy metals have toxic effects on fish. Insufficient measures are a serious problem in our country and around the world. This problem can threaten human health in areas where it is common for people to obtain nutrition from local bodies of water. In this study, the toxicological impacts of some heavy metals were investigated on carbonic anhydrase activity in gilthead gills. Carbonic anhydrase (CA) was purified from gilthead sea bream (Sparus aurata) gills with a specific activity of 2872.92EUmg-1 and a yield of 32.84% using affinity chromatography. The overall purification was approximately ~84-fold. SDS-polyacrylamide gel electrophoresis showed a single band, and the MW was approximately 30.5kDa (Soyut et al., 2008, 2012; Soyut and Beydemir, 2008, 2012; Kaya et al., 2013). The kinetic and characteristic properties of CA such as the optimum pH, stable pH, optimum temperature, activation energy (Ea), activation enthalpy (?H), Q10, Km and Vmax were determined. Cadmium (Cd2+), copper (Cu2+), nickel (Ni2+) and silver (Ag+) inhibited CA activity in in vitro conditions. Ki values were calculated for these metals. Ki values were 31.20mM for cadmium (Cd2+), 161.96mM for copper (Cu2+), 10.79mM for nickel (Ni2+) and 0.0082mM for silver (Ag+) based on Lineweaver-Burk plots. Except for cadmium, heavy metals had the same inhibition mechanism. Cadmium was competitive, and the others were noncompetitive. © 2015 Elsevier B.V.
