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Biological systems respond appropriately to a variety of environments thus representing complex systems with rich physiological behaviour. Quantitative models can be used to identify the design components that result in the system complexity. In this work, the tryptophan system of Escherichia coli that synthesises tryptophan internally when faced with starvation in a rapid manner and shuts off the synthesis sluggishly when the cells are exposed to a medium replete with tryptophan has been discussed. The evolved regulatory design is capable of providing such an asymmetric response that represents an appropriate behaviour to ensure survival. The tryptophan system uses three distinct regulatory mechanisms namely genetic regulation, transcriptional attenuation and enzyme inhibition to achieve its goals. It has been shown that genetic repression and attenuation are the only active regulatory mechanisms during moderate and severe starvation. However, as the degree of starvation increases, repression is relieved prior to attenuation. The analysis also shows that enzyme inhibition does not play a role under severe starvation and plays a marginal role in increasing the rate of repression when the cells are exposed to well-fed conditions. Finally, we use tools from linear systems theory to rationalise the above observations based on the poles and zeros of an approximated linear system.

Biological networks have evolved through adaptation in uncertain environments. Of the different possible design paradigms, some may offer functional advantages over others. These designs can be quantified by the structure of the network resulting from molecular interactions and the parameter values. One may, therefore, like to identify the design motif present in the evolved network that makes it preferable over other alternatives. In this work, we focus on the regulatory networks characterized by serially arranged processes, which are regulated by multiple feedback loops. Specifically, we consider the tryptophan system present in Escherichia coli, which may be conceptualized as three processes in series, namely transcription, translation and tryptophan synthesis. The multiple feedback loop motif results from three distinct negative feedback loops, namely genetic repression, mRNA attenuation and enzyme inhibition. A framework is introduced to identify the key design components of this network responsible for its physiological performance. We demonstrate that the multiple feedback loop motif, as seen in the tryptophan system, enables robust performance to variations in system parameters while maintaining a rapid response to achieve homeostasis. Superior performance, if arising from a design principle, is intrinsic and, therefore, inherent to any similarly designed system, either natural or engineered. An experimental engineering implementation of the multiple feedback loop design on a two-tank system supports the generality of the robust attributes offered by the design.

Living systems must adapt quickly and stably to uncertain environments. A common theme in cellular regulation is the presence of multiple feedback loops in the network. An example of such a feedback structure is regulation of tryptophan concentration in Escherichia coli. Here, three distinct feedback mechanisms, namely genetic regulation, mRNA attenuation and enzyme inhibition, regulate tryptophan synthesis. A pertinent question is whether such multiple feedback loops are "a case of regulatory overkill, or do these different feedback regulators have distinct functions?" Another moot question is how robustness to uncertainties can be achieved structurally through biological interactions. Correlation between the feedback structure and robustness can be systematically studied by tools commonly employed in feedback theory. An analysis of feedback strategies in the tryptophan system in E. coli reveals that the network complexity arising due to the distributed feedback structure is responsible for the rapid and stable response observed even in the presence of system uncertainties.

A mathematical model has been developed to study the effect of external tryptophan on the trp operon. The model accounts for the effect of feedback repression by tryptophan through the Hill equation. We demonstrate that the trp operon maintains an intracellular steady-state concentration in a fivefold range irrespective of extracellular conditions. Dynamic behavior of the trp operon corresponding to varying levels of extracellular tryptophan illustrates the adaptive nature of regulation. Depending on the external tryptophan level in the medium, the transient response ranges from a rapid and underdamped to a sluggish and highly overdamped response. To test model fidelity, simulation results are compared with experimental data available in the literature. We further demonstrate the significance of the biological structure of the operon on the overall performance. Our analysis suggests that the tryptophan operon has evolved to a truly optimal design.