Bacteria contain sigma (sigma) factors that control gene expression in response to various environmental stimuli. The alternative sigma factors sigma(FPVI) and sigma(FPDS) bind specifically to the antisigma factor FpvR. These proteins are an essential component of the pyoverdine-based system for iron uptake in Pseudomonas aeruginosa. Due to the uniqueness of this system, where the activities of both the sigma(FPVI) and sigma(PVDS) sigma factors are regulated by the same antisigma factor, the interactions between the antisigma protein FpvR(20) and the sigma(FPVI) and sigma(FPDS) proteins have been widely studied in vivo. However, difficulties in obtaining soluble, recombinant preparations of the sigma(FPVI) and sigma(FPDS) proteins have limited their biochemical and structural characterizations. In this study, we describe a purification protocol that resulted in the production of soluble, recombinant His(6)-sigma(FPVI)/FpvR(1-67), His(6)-sigma(FPVI)/FpVR(1-89), His(6)-sigma(PVDS)/FpvR(1-67) and His(6)-sigma(PVDS)/FPvR(1-89) protein complexes (where FpvR(1-67) and FpvR(1-89) are truncated versions of FpvR(20)) at high purities and concentrations, appropriate for biophysical analyses by circular dichroism spectroscopy and analytical ultracentrifugation. These results showed the proteins to be folded in solution and led to the determination of the affinities of the protein-protein interactions within the His(6)-sigma(FPVI)/ FpvR(1-67) and His(6)-sigma(PVDS)/EpvR(1-67) complexes. A comparison of these values with those previously reported for the His(6)-sigma(FPVI)/FpvR(1-89) and His(6)-sigma(PVDS)/FpvR(1-89) complexes is made.