N5-59, a siderophore deficient mutant strain of Rhodococcus erythropolis strain IGTS8 (IGTS8) was investigated to learn more about how this poorly characterized bacterium acquires iron. N5-59 cells were starved for iron and then lysed to release any intracellular siderophore. No intracellular siderophore was detected indicating that N5-59 is not defective in the export or release of siderophore but is probably unable to synthesize siderophore. In a cross-feeding bioassay, growth of N5-59 (in an iron depleted medium) was greatly enhanced by the addition of exogenous siderophore from IGTS8 and other Rhodococcus species indicating that N5-59 is not defective in siderophore uptake. A DNA hybridization probe was prepared using DNA flanking the site of insertional mutation in strain N5-59. This probe was then used to clone a 6 kilobase pair, PstI restriction fragment from the chromosome IGTS8. This cloned DNA is expected to contain the intact gene(s) that was interrupted in N5-59.

Iron is an essential nutrient for most bacteria because enzymes like nitrate reductase and cytochromes use it as a cofactor. However, in most aerobic, neutral pH environments, iron is essentially insoluble and not easily available for bacteria to use. Many bacteria respond to this problem by releasing small organic compounds called siderophores that bind and effectively solubilize iron so that it can be transported into the cell for growth. The focus of this study was to learn more about the iron acquisition and especially the transport of iron by the soil bacterium Rhodococcus erythropolis. To fulfill this aim, mutant strains of the bacteria were screened for those that overproduce siderophore. Often, a bacterium will over produce siderophore to compensate for a defect in transport. One such mutant, R187-12, was further analyzed by cloning the region of the chromosome containing the defective gene responsible for over production of siderophore into a plasmid vector. The DNA sequence of this region was determined and analyzed for the presence of similar genes encoding transport proteins.