Inactivation of Glucose-6-Phosphate Dehydrogenase in Solution by Low- and High-Frequency Ultrasound

• Published in: Applied biochemistry and microbiology 2004-03, Vol.40 (2), p.120-128
• Main Authors: Rachinskaya, Zh. V., Karasyova, E. I., Metelitza, D. I.
• Format: Article
• Language: English
• Subjects: Dehydrogenase; Ethanol; Free radicals; Inactivation; Kinetics
• ISSN: 0003-6838; 1608-3024
• DOI: 10.1023/B:ABIM.0000018913.59738.9a

We compared the kinetics of glucose-6-phosphate dehydrogenase (G6PDH, EC 1.1.1.49) inactivation in 0.1 M phosphate buffer (pH 7.4) at 36-50°Ð¡ under conditions of exposure to low-frequency (LF, 27 kHz, 60 W/cm^sup 2^) or high-frequency (HF, 880 kHz, 1.0 W/cm^sup 2^) ultrasound (USD). The inactivation of G6PDH was characterized by effective first-order rate constants: k^sub in^, total inactivation; k^sub in^^sup *^, thermal inactivation; and k^sub in^(usd), ultrasonic inactivation. Dilution of the enzyme solution from 20 to 3 nM was accompanied by a significant increase in the values of the three rate constants. The following inequality was valid in all cases: k^sub in^ > k^sub in^^sup *^. The rate constants increased with temperature. The Arrhenius plots of the temperature dependences of k^sub in^ and k^sub in^(usd) had an break point at 44°C. The activation energy (Ð*^sub act^) of the total inactivation of G6PDH was higher than Ð*^sub act^ for the process of ultrasonic inactivation of this enzyme. The two values were found to depend on USD frequency: Ð*^sub act^ was higher in the case of inactivation with low-frequency ultrasound (LF-USD) than high-frequency ultrasound (HF-USD). The rate of the ultrasonic inactivation of this enzyme substantially decreased in the presence of low concentrations of HO^sup .^ radical scavengers (dimethylformamide, ethanol, and mannitol). This fact supports the conclusion that free radicals are involved in the mechanism of G6PDH inactivation in solutions exposed to LF-USD and HF-USD. Ethanol was an effective protector of G6PDH inactivation in solutions exposed to USD.[PUBLICATION ABSTRACT]