The effect of chemical (urea) and physical (temperature and high pressure) denaturation on the structural properties of soybean lipoxygenase-1 (LOX1) was analyzed through dynamic fluorescence spectroscopy and circular dichroism. We show that the fluorescence decay of the native protein could be fitted by two lorentzian distributions of lifetimes, centered at 1 and 4 ns. The analysis of the urea-denatured protein suggested that the shorter distribution is mostly due to the tryptophan residues located in the N-terminal domain of LOX1. We also show that a pressure of 2400 bar and a temperature of 55degreesC brought LOX-1 to a state similar to a recently described stable intermediate "I." Analysis of circular dichroism spectra indicated a substantial decrease of alpha-helix compared with beta-structure under denaturing conditions, suggesting a higher stability of the N-terminal compared with the C-terminal domain in the denaturation process. (C) 2001 Elsevier Science.
Effect of denaturants on the structural properties of soybean lipoxygenase-1
Maccarrone M
2001-01-01
Abstract
The effect of chemical (urea) and physical (temperature and high pressure) denaturation on the structural properties of soybean lipoxygenase-1 (LOX1) was analyzed through dynamic fluorescence spectroscopy and circular dichroism. We show that the fluorescence decay of the native protein could be fitted by two lorentzian distributions of lifetimes, centered at 1 and 4 ns. The analysis of the urea-denatured protein suggested that the shorter distribution is mostly due to the tryptophan residues located in the N-terminal domain of LOX1. We also show that a pressure of 2400 bar and a temperature of 55degreesC brought LOX-1 to a state similar to a recently described stable intermediate "I." Analysis of circular dichroism spectra indicated a substantial decrease of alpha-helix compared with beta-structure under denaturing conditions, suggesting a higher stability of the N-terminal compared with the C-terminal domain in the denaturation process. (C) 2001 Elsevier Science.Pubblicazioni consigliate
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