Changes in carbonic anhydrase activity and gene expression of Hsp70 in rainbow trout (Oncorhynchus mykiss) muscle after exposure to some metals

Authors : Hakan SÖYÜT, Şükrü BEYDEMİR, Saltuk Buğrahan CEYHUN

Pages : 499-508

View : 21 | Download : 9

Publication Date : 0000-00-00

Article Type : Research Paper

Abstract :The effects of some heavy metals Coinsert ignore into journalissuearticles values(II);, Cuinsert ignore into journalissuearticles values(II);, Zninsert ignore into journalissuearticles values(II);, and Aginsert ignore into journalissuearticles values(I); on carbonic anhydrase insert ignore into journalissuearticles values(CA); activity from rainbow trout insert ignore into journalissuearticles values(RT); muscle were investigated. Moreover, the effects of these metals on the expression of heat shock protein 70 insert ignore into journalissuearticles values(Hsp70); gene in muscle tissue were examined by real-time quantitative PCR insert ignore into journalissuearticles values(RT-PCR); in muscle tissue after 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 insert ignore into journalissuearticles values(Mw); of subunit and native enzyme were approximately 30 and 31 kDa, respectively. Optimum pH, stable pH, optimum temperature, activation energy insert ignore into journalissuearticles values(Ea);, activation enthalpy insert ignore into journalissuearticles values(DH);, and Q10 insert ignore into journalissuearticles values(the difference in activity of enzyme caused by the increment of 10 °C); value were determined. Apparent Michaelis constant insert ignore into journalissuearticles values(Km);, maximum reaction rate insert ignore into journalissuearticles values(Vmax);, and turnover rate of the enzyme insert ignore into journalissuearticles values(Kcat); values were 1.29 mM, 0.17 µmol min-1, and 28.8 s-1, respectively. The catalytic efficiency insert ignore into journalissuearticles values(Kcat/Km); was 22.3. The heavy metals decreased in vitro CA activity. Inhibition mechanisms of the metal ions were noncompetitive, except for the Coinsert ignore into journalissuearticles values(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 Coinsert ignore into journalissuearticles values(II); > Zninsert ignore into journalissuearticles values(II); > Cuinsert ignore into journalissuearticles values(II); > Aginsert ignore into journalissuearticles values(I);. Interestingly, Aginsert ignore into journalissuearticles values(I); was the most effective metal ion on Hsp70 gene expression.
Keywords : Key words Carbonic anhydrase, characterization, muscle, rainbow trout, heavy metals, heat shock protein 70