As the Limiting substrate is altered, a microorganism’s internal structure is also altered by invoking different metabolic pathways for the utilization of the limiting substrate. Transitions between pathways are moderated by the processes of metabolic regulation and are observed under steady-state and transient conditions. Under nitrogen-limited steady-state conditions, the effects of overflow metabolism are observed in the form of polysaccharide synthesis and excess carbon oxidation, two phenomena that are not observed under carbon-limited conditions. Furthermore, during transient periods following dilution rate shifts, metabolic lags are observed to be a function of the size of the shift as well as the limiting substrate. The latter observation indicates the preference of one reaction pathway over another as the status of glucose or NH4+ is altered from limiting to nonlimiting. This paper presents steady-state and transient results from continuous culture experiments using Escherichia coli W. Singly-Limiting conditions, when either glucose or ammonia is the limiting substrate, are investigated. Transient conditions are created by quickly increasing or decreasing the dilution rate of the fermenter. By utilizing the control strategies identified by Straight and Ramkrishna (1994) for regulating the processes commonly found within metabolic pathways, a cybernetic model is developed and compared to the steady-state and transient experimental results. Due to the incorporation of metabolic pathways, the development of the model accounts for lumped biosynthetic intermediates in addition to key enzymes that catalyze different cellular processes. The model also accounts for an internal resource that is optimally allocated toward the synthesis of the key enzymes. Furthermore, the model incorporates the effects of maintenance processes and overflow metabolism. Upon incorporating nitrogen utilization, the kinetic aspects of the model do not explicitly reduce to those of previous cybernetic models; however, the regulatory structure is in complete agreement with previous cybernetic models proposed for carbon utilization.