Changes in the iron coordination sphere of Fe(II) lipoxygenase-1 from soybeans upon binding of linoleate or oleate.

Fe K-edge X-ray absorption spectra of the non-heme iron constituent of lipoxygenase-1 from soybeans were obtained. The spectrum of 2.5 mM Fe(II) lipoxygenase, mixed with 1.2 M linoleate in the absence of O2, was compared to the spectrum of the native (i.e. untreated) enzyme. In the lipoxygenase-linoleate complex, an edge shift to lower energy was observed. This indicated that the iron-ligand distances in this complex are slightly longer than those in the untreated enzyme species. The extended X-ray absorption fine structure spectrum of Fe(II) lipoxygenase, prepared by anaerobic reduction of 2.5 mM Fe(II) lipoxygenase with 1.2 M linoleate, was very similar to the spectrum of the anaerobic lipoxygenase-linoleate complex. We conclude that the conformational differences between the iron coordination spheres of native and cycled Fe(II) lipoxygenase must be ascribed to the presence of linoleate, and not to changes in the enzyme that occur only after one cycle of oxidation and reduction. Furthermore, spectra of 2.5 mM Fe(II) lipoxygenase mixed with 1.2 M oleate, either in the absence or in the presence of O2, were also identical to the spectrum of the Fe(II) lipoxygenase-linoleate complex. This finding is in agreement with our observation that oleate is a competitive inhibitor of the lipoxygenase reaction. Moreover, the similarity of the lipoxygenase-oleate complexes in the presence and absence of O2 excludes the possibility that O2 binding to the iron cofactor is induced upon binding of a fatty acid to lipoxygenase.