Effect of denaturants on the structural properties of soybean lipoxygenase-1.
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 55 degrees C 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.
Malvezzi-Campeggi
F
F
INFM, University of Rome, Tor Vergata, Rome, I-00133, Italy.
Rosato
N
N
Finazzi-Agrò
A
A
Maccarrone
M
M
D002942:Circular Dichroism; D004795:Enzyme Stability; D006358:Hot Temperature; D008084:Lipoxygenase / Q000737:chemistry*; D011312:Pressure; D011489:Protein Denaturation / Q000187:drug effects; D017434:Protein Structure, Tertiary; D013025:Glycine max / Q000201:enzymology*; D013050:Spectrometry, Fluorescence; D014364:Tryptophan / Q000737:chemistry; D014508:Urea / Q000494:pharmacology
D014364:Tryptophan; D014508:Urea; C101084:lipoxygenase L-1; D008084:Lipoxygenase