Bengal gram lipoxygenase: fluorescence quenching study of the interaction of linoleic acid and 13- and 9-hydroperoxylinoleic acids with the two forms of the enzyme

The intrinsic protein fluorescence of the two forms of lipoxygenases from Bengal gram has been characterized. The
fluorescence is dominated by emission from tryptophan residues in a hydrophobic environment. The substrate linoleic
acid and the reaction products 13- and 9-hydroperoxylinoleic acids quenched the intrinsic protein fluorescence equally
for two forms of the enzyme without lag period. From the fluorescence quenching measurements, the association
constant (K) and the free energy change for the interaction have been calculated. The two forms of the enzyme differ
in their affinity to the substrate. The ΔG° value for the interaction of substrate/products was calculated to be -5.0 kcal/
mol, suggesting that the interaction is a weak one. Spectroscopic measurements do not indicate a large conformational
change in the enzyme due to the binding of these molecules.