The functional principle of eukaryotic molybdenum insertases.

The molybdenum cofactor (Moco) is a redox-active prosthetic group found in the active site of Moco- dependent enzymes, which are vitally important for life. Moco biosynthesis involves several enzymes that catalyze the subsequent conversion of GTP to cyclic pyranopterin monophoshpate (cPMP), molybdopterin (MPT), adenylated MPT (MPT-AMP) and finally Moco. While the underlying principles of cPMP, MPT, and MPT-AMP formation are well-understood, the molybdenum insertase (Mo-insertase) catalyzed final Moco maturation step is not. In this study we analyzed high resolution X-ray data sets of the plant Mo-insertase Cnx1E that revealed two molybdate binding sites within the active site, hence improving the current view on Cnx1E functionality. The presence of molybdate anions in either of these sites is tied to a distinctive backbone conformation, which we suggest to be essential for Mo-insertase molybdate selectivity and insertion efficiency.