Stochastic kinetic analysis of the Escherichia coli stress circuit using sigma(32)-targeted antisense

A stochastic Petri net model was developed for simulating the sigma (32) st ress circuit In E. coli. Transcription factor sigma (32) is the principal r egulator of the response of E. coli to heat shock. Stochastic Petri net (SP N) models are well suited for kinetics characterization of fluxes in bioche mical pathways. Notably, there exists a one-to-one mapping of model tokens and places to molecules of particular species. Our model was validated agai nst experiments in which ethanol (inducer of heat shock response)and sigma (32)-targeted antisense (downward regulator) were used to perturb the sigma (32) regulatory pathway. The model was also extended to simulate the effec ts of recombinant protein production. Results show that the stress response depends heavily on the partitioning of sigma (32) within the cell; that is , sigma (32) becomes immediately available to mediate a stress response bec ause it exists primarily in a sequestered, inactive form, complexed with ch aperones DnaK, DnaJ, and GrpE. Recombinant proteins, however, also compete for chaperone proteins, particularly when folded Improperly. Our simulation s indicate that when the expression of recombinant protein has a low requir ement for DnaK, DnaJ, and GrpE, the overall sigma (32) levels may drop, but the level of heat shock proteins will increase. Conversely, when the overe xpressed recombinant protein has a strong requirement for the chaperones, a severe response is predicted. Interestingly, both cases were observed expe rimentally. (C) 2001 John Wiley & Sons, Inc.