Spectroscopic Characterization of the Isolated Iron–Molybdenum Cofactor (FeMoco) Precursor from the Protein NifEN

Abstract

The FeMoco of Mo-nitrogenase provides the active site for substrate reduction. Previously, a FeMoco precursor was captured on NifEN, a scaffold protein for FeMoco biosynthesis. Here, we report the isolation of FeMoco precursor from NifEN. The integrity of the precursor is reflected by the restoration of the precursor-specific EPR signal, as well as the full proficiency of the precursor in biosynthesis and catalysis upon its incorporation into the precursor-deficient NifEN. Moreover, XAS/EXAFS analysis supports the eight-iron model of the precursor, suggesting that the insertion of heterometal into the precursor involves exchanging one terminal iron atom for a molybdenum atom.