There are few safe and effective drugs available that protect healthy tissue against radiation-induced damage, highlighting the need to develop such radioprotective agents. We investigated the mechanism underlying the protective effects of the novel, recombinant, flagellin-like protein FlaA N/C against radiation-induced tissue damage in female BALB/c mice. FlaA N/C treatment increased the levels of several pro-proliferative cytokines while inhibiting apoptosis and reducing inflammation. These effects were accompanied by activation of the nuclear factor κB signaling pathway via direct interaction of FlaA N/C with Toll-like receptor 5, as well as enhanced survival of mice after total-body irradiation compared to that observed with treatment with amifostine, a radioprotective agent that is currently being used in clinical practice. These results indicate that FlaA N/C could be further explored as a possible protector of damage to healthy tissue during radiotherapy.