Dpr is an iron-binding protein required for oxygen tolerance in Streptococcus mutans. We previously proposed that Dpr could confer oxygen tolerance to the bacterium by sequestering intracellular free iron ions that catalyze generation of highly toxic radicals (Y. Yamamoto, M. Higuchi, L. B. Poole, and Y. Kamio, J. Bacteriol. 182:3740-3747, 2000; Y. Yamamoto, L. B. Poole, R. R. Hantgan, and Y. Kamio, J. Bacteriol. 184:2931-2939, 2002). Here, we examined the intracellular free iron status of wild-type (WT) and dpr mutant strains of S. mutans, before and after exposure to air, by using electron spin resonance spectrometry. Under anaerobic conditions, free iron ion concentrations of WT and dpr strains were 225.9 +/- 2.6 and 333.0 +/- 61.3 microM, respectively. Exposure of WT cells to air for 1 h induced Dpr expression and reduced intracellular free iron ion concentrations to 22.5 +/- 5.3 microM; under these conditions, dpr mutant cells maintained intracellular iron concentration at 230.3 +/- 28.8 microM. A decrease in cell viability and genomic DNA degradation was observed in the dpr mutant exposed to air. These data indicate that regulation of the intracellular free iron pool by Dpr is required for oxygen tolerance in S. mutans.