[Spectral analysis of polyphenol oxidase (PPO) and lipoxygenase (LOX) treated by pulsed electric field].
Inactivation effect of pulsed electric field (PEF) on polyphenol oxidase (PPO) and lipoxygenase (LOX) was investigated using a laboratory PEF system with a coaxial treatment chamber. Circular dichroism (CD) and fluorescence analysis were used to study the conformation change of the protein. The experimental results show that PPO and LOX can be effectively inactivated by the PEF treatment. Inactivation effect of PPO and LOX increases with the increase in the applied electric strength and the treatment time. Activity of PPO and LOX can be reduced by 60.3% and 21.7% at 20 kV x cm(-1) after being treated for 320 micros respectively. The decrease of the negative peaks (208 and 215 nm in PPO spectra, 208 nm and 218 nm in LOX spectra) in CD spectra of PPO and LOX shows that PEF treatment caused a loss of alpha-helix and increase in beta-sheet content, indicating that conformation changes occur in the secondary structure of PPO and LOX enzyme. This effect was strengthened as the applied electric field increased: alpha-helical content of PPO and LOX was 56% and 29% after being treated at 8 kV x cm(-1), however, when the electric field was increased up to 20 kV x cm(-1), alpha-helical content of PPO and LOX decreased to 21% and 16% respectively. The decrease rate of alpha-helix and increase rate of beta-sheet in PPO are higher than LOX, indicating that the second conformation of PPO is less resistant to PEF treatment than LOX. The fluorescence intensity of LOX increases after PEF treatment. At the same time, increasing the applied pulsed electric field increases the fluorescence intensity emitted. Fluorescence measurements confirm that tertiary conformation changes occur in the local structure of LOX. However the possible mechanism of the conformation change induced by the PEF treatment is beyond the scope of the present investigation.
Luo
Wei
W
Lab of New Technologies for Energy and Electrical Engineering, Shenzhen Graduate School, Tsinghua University, Shenzhen 518055, China.
Zhang
Ruo-Bing
RB
Chen
Jie
J
Wang
Li-Ming
LM
Guan
Zhi-Cheng
ZC
Jia
Zhi-Dong
ZD
D004156:Catechol Oxidase / Q000737:chemistry*; D002942:Circular Dichroism; D004560:Electricity; D005453:Fluorescence; D008084:Lipoxygenase / Q000737:chemistry*; D017433:Protein Structure, Secondary
D004156:Catechol Oxidase; D008084:Lipoxygenase