The goal of this research is an understanding, at a biochemical level, of the steps necessary to make a functional nitrogenase complex. This goal will be achieved by a biochemical and genetic analysis of the nif regulon of Klebsiella pneumoniae. Mutants will be generated and genetically characterized with defects in """"""""nonessential-nif genes. Conditions will be found where such mutants display a Nif-phenotype. The functions of the products of six nif genes is presently unknown and will be determined using a variety of assays which determine (1) the integrity of electron transport to nitrogenase, (2) the electrophoretic properties of components of the nitrogenase system, (3) the presence of iron-sulphur centers in nif-related proteins, (4) the cellular location of nif proteins. Since molybdenum is of central importance to nitrogenase, mutants defective in molybdenum metabolism will be isolated and characterized genetically and biochemically. Such information will be applicable to the understanding of molybdenum metabolism in other enzyme systems and in other organisms less amenable to genetic analysis.
