Transgenic bacteria producing pyrroloquinoline quinone, a known cofactor for dehydrogenases and an inducer of a periplasmic protein kinase activity, show resistance to both oxidative stress and protection from nonoxidative effects of radiation and DNA-damaging agents. Deinococcus radiodurans R1 encodes an active pyrroloquinoline quinone synthase, and constitutive synthesis of pyrroloquinoline quinone occurred in wild-type bacteria. Disruption of a genomic copy of pqqE resulted in cells that lacked this cofactor. The mutant showed a nearly 3-log decrease in gamma radiation resistance and a 2-log decrease in mitomycin C tolerance compared to wild-type cells. The mutant cells did not show sensitivity to UVC radiation. Expression of pyrroloquinoline quinone synthase in trans showed that there was functional complementation of gamma resistance and mitomycin C tolerance in the pqqE mutant. The sensitivity to gamma radiation was due to impairment or slow kinetics of DNA double strand break repair. Low levels of (32)P incorporation were observed in total soluble proteins of mutant cells compared to the wild type. The results suggest that pyrroloquinoline quinone has a regulatory role as a cofactor for dehydrogenases and an inducer of selected protein kinase activity in radiation resistance and DNA strand break repair in a radioresistant bacterium.