Internal pool sizes of glutamine and glutamate in Klebsiella pneumoniae grown under nitrogen limitation or nitrogen sufficiency were measured to study the signal transduction of external nitrogen limitation. K pneumoniae cells were grown in an anaerobic, ammonium-limited chemostat culture. At a growth rate of 0.217 h(-1), the steady state ammonium concentration in the culture was 55 muM, correlating with repression of the nitrogen fixation (nif) genes. At growth rates below 0.138 h(-1), the ammonium concentration in the culture dropped below 0.5 muM and the nif genes became derepressed. During the transition from nitrogen sufficiency to nitrogen limitation, the internal glutamine pool in K. pneumoniae decreased by a factor of approximately 6. The glutamate pool, however, remained stable. Similarly, in anaerobic batch cultures with different limiting nitrogen sources, the glutamine pool generally decreased by a factor of 7 to 9 when nif gene derepression was achieved. All the limiting nitrogen sources used resulted in decreased growth rates compared with growth under nitrogen excess, suggesting an inverse relationship between glutamine pool size and doubling time. These studies indicate that K. pneumoniae perceives external nitrogen limitation as internal glutamine limitation.