REGULATORY PROCESSES IN ULTRA SLOW GROWING MICROORGANISMS (ESCHERICHIA COLI, BACILLUS POLYMYXA)
In a continuous fermentor with 100% recycle, growth of Escherichia coli and Bacillus polymyxa occurs in three sharply distinct phases: an initial phase of exponential growth that terminates when the glucose level in the fermentor falls below transport saturation levels; a second phase of linear growth whose rate is dependent on the rate of glucose provision to the fermentor, but whose length if fixed and independent of growth rate; and a third phase of slower glucose dependent linear growth. The specific growth rate falls in phase 2 and 3 while the mass doubling time lengthen, eventually reaching hundreds of hours. Guanosine 5-diphosphate 3-diphosphate (ppGpp) accumulation commences at the beginning of phase 2 in E. coli, reaching a maximum at the start of phase 3, concurrent with the slowing of RNA synthesis. This final growth stage then is an indefinitely prolonged state of regulation by the stringent response. The length of growth phase 2 was changed substantially by mutations in the relA, relX, and spoT loci, in ways which are exactly predictable from their effects on ppGpp synthesis, as phase 2 was longer for mutants with a decreased capability for ppGpp synthesis, and shorter for mutants with a decreased capability for ppGpp degradation. In B. polymyxa, the cost of the stringent response in phase 3 is approximately 9% of the available energy, while the cost of extracellular anabolites ranges from 8 to 11% of the available energy. Both are maintenance energy costs by definition. After a carbon upshift in phase 3, the population growth rate and ppGpp concentration in B. polymyxa and E. coli proceeds to a level seen in phase 2. Fermentor levels of cAMP in E. coli rise abruptly at each phase transition, then decline throughout the phases. The inducible DNA repair system, "SOS", can be activated in phase 3 cultures of E. coli. The degradation or turnover of proteins involved with this system, is apparently very slow, if compared with the rates of degradation of the inducible enzymes, B-galactosidase and tryptophanase. The activity of this system in phase 3 cells is interpretable as signifying the presence of a chromosome replicating fork on only 38% of the population.
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