Pseudomonas aeruginosa strain CSU, a nongenetically engineered bacterial strain known to bind dissolved hexavalent uranium (as UO22+ and/or its cationic hydroxo complexes), was characterized with respect to its sorptive activity (equilibrium and dynamics). Living, heat-killed, permeabilized, and unreconstituted lyophilized cells were all capable of binding uranium. The uranium biosorption equilibrium could be described by the Langmuir isotherm. The rate of uranium adsorption increased following permeabilization of the outer and/or cytoplasmic membrane by organic solvents such as acetone. P. aeruginosa CSU biomass was significantly more sorptive toward uranium than certain novel, patented biosorbents derived from algal or fungal biomass sources. P. aeruginosa CSU biomass was also competitive with commercial cation-exchange resins, particularly in the presence of dissolved transition metals. Uranium binding by P. aeruginosa CSU was clearly pH dependent. Uranium loading capacity increased with increasing pH under acidic conditions, presumably as a function of uranium speciation and due to the H+ competition at some binding sites. Nevertheless, preliminary evidence suggests that this microorganism is also capable of binding anionic hexavalent uranium complexes. Ferric iron was a strong inhibitor of uranium binding to P. aeruginosa CSU biomass, and the presence of uranium also decreased the Fe3+ loading when the biomass was not saturated with Fe3+, suggesting that Fe3+ and uranium may share the same binding sites on biomass. Although the equilibrium loading capacity of uranium was greater than that of Fe3+, this biomass showed preference of binding Fe3+ over uranium. Thus, a two-stage process in which iron and uranium are removed in consecutive steps was proposed for efficient use of the biomass as a biosorbent in uranium removal from mine wastewater, especially acidic leachates.