INVOLVEMENT OF THE NARJ AND MOB GENE-PRODUCTS IN DISTINCT STEPS IN THE BIOSYNTHESIS OF THE MOLYBDOENZYME NITRATE REDUCTASE IN ESCHERICHIA-COLI

The Escherichia coil mob locus is required for synthesis of active mol ybdenum cofactor, molybdopterin guanine dinucleotide. The mobB gene is not essential for molybdenum cofactor biosynthesis because a deletion of both mob genes can be fully complemented by just mobA. Inactive ni trate reductase, purified from a mob strain, can be activated in vitro by incubation with protein FA (the mobA gene product), GTP, MgCl2, an d a further protein fraction, factor X. Factor X activity is present i n strains that lack MobB, indicating that it is not an essential compo nent of factor X, but overexpression of MobB increases the level of fa ctor X, MobB, therefore, can participate in nitrate reductase activati on. The narJ protein is not a component of mature nitrate reductase bu t narJ mutants cannot express active nitrate reductase A. Extracts fro m narJ strains are unable to support the in vitro activation of purifi ed mob nitrate reductase: they lack factor X activity. Although the mo b gene products are necessary for the biosynthesis of all E. coli moly bdoenzymes as a result of their requirement for molybdopterin guanine dinucleotide, NarJ action is specific for nitrate reductase A. The ina ctive nitrate reductase A derivative in a narJ strain can be activated in vitro following incubation with cell extracts containing the narJ protein, NarJ acts to activate nitrate reductase after molybdenum cofa ctor biosynthesis is complete.