Ukr.Biochem.J. 2015; Volume 87, Issue 1, Jan-Feb, pp. 75-82

doi: https://doi.org/10.15407/ubj87.01.075

Analysis of creatine kinase activity with evaluation of protein expression under the effect of heat and hydrogen peroxide

20.05.2015   Uncategorized   No comments

A. D. Rakhmetov1, Lee Sang Pil2, L. I. Ostapchenko1, Chae Ho Zoon2

1Education and Science Center Institute of Biology Taras Shevchenko National University of Kyiv, Ukraine;
2Chonnam National University, Gwangju, South Korea;
e-mail: anar.rakhmetov@gmail.com

Protein oxidation has detrimental effects on the brain functioning, which involves inhibition of the crucial enzyme, brain type creatine kinase (CKBB), responsible for the CK/phosphocreatine shuttle system. Here we demonstrate a susceptibility of CKBB to several ordinary stressors. In our study enzymatic activity of purified recombinant brain-type creatine kinase was evaluated. We assayed 30 nM concentration of CKBB under normal and stress conditions. In the direction of phosphocreatine formation hydrogen peroxide and heat treatments altered CKBB activity down to 26 and 14%, respectively. Also, examination of immunoblotted membrane patterns by SDS-PAGE electrophoresis and western blot analysis showed a decrease in expression levels of intrinsic CKBB enzyme in HeLa and A549 cells. Hence, our results clearly show that cytosolic CKBB is extremely sensitive to oxidative stress and heat induced inactivation. Therefore, due to its susceptibility, this enzyme may be defined as a potential target in brain damage.

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