MOLYBDENUM COFACTOR BIOSYNTHESIS - THE PLANT PROTEIN CNX1 BINDS MOLYBDOPTERIN WITH HIGH-AFFINITY

The molybdenum cofactor is an essential part of all eukaryotic molybdo enzymes. It is a molybdopterin (RIFT) revealing the same core structur e in all organisms, The plant protein Cnx1 from Arabidopsis thaliana i s involved in the multi step biosynthesis of molybdenum cofactor, Prev ious studies (Stallmeyer, B., Nerlich, A., Schiemann, J., Brinkmann, H ., and Mendel, R, R, (1995) Plant J, 8, 751-762) suggested a function of Cnx1 in a late step of cofactor biosynthesis distal to the formatio n of MPT, i.e, conversion of MPT into molybdenum cofactor, Here we pre sent the first biochemical evidences confirming this assumption, The p rotein Cnx1 consists of two domains (E and G) homologous to two distin ct Escherichia coli proteins involved in cofactor synthesis. Binding s tudies with recombinantly expressed and purified Cnx1 and with its sin gle domains revealed a high affinity of the G domain to MPT (kD = 0.1 mu M) with equimolar binding, In contrast, the E domain of Cnx1 binds MPT with lower affinity (kD = 1.6 mu M) and in a cooperative manner (n (H) = 1.5), The entire Cnx1 showed a tight and cooperative MPT binding . Based on these data providing a common link between both domains tha t matches the previous characterization of plant and bacterial Cnx1 ho mologous mutants, we present a model for the function of Cnx1.