Urease expression in Streptococcus salivarius 57.1 is induced by acidic pH, and further enhanced at high growth rate and with excess carbohydrate. Notably, the phosphoenolpyruvate (PEP):sugar phosphotransferase system (PTS) activity is repressed in oral streptococci under the same conditions. To test the hypothesis that the PTS may be involved in urease regulation, spontaneous mutants (PTS-1 and PTS-4) that were resistant to 2-deoxyglucose were isolated. When compared to wild-type, PTS-1 was devoid of enzyme IIAManH (EIIAManII) and synthesized low amounts of EIIAManL, and PTS-4 was devoid of EIIAManL, but produced wild-type levels of EIIAManH. Urease expression was examined in continuous chemostat cultures at steady state. Induction by acidic pH was still observed in both mutants, but at lower levels compared to wild-type, under carbohydrate limiting conditions. Conversely, the lower level of expression in PTS-4 could be overcome in excess carbohydrate. The data indicated evidence of a molecular link between the PTS, sugar metabolism, and regulation of urease expression.