NOT Gates Based on Protein Degradation as a Case Study for a New Modular Modeling via SBML Level 3-Comp Package

In 2008, we were among the first to propose a method for the visual design and modular modeling of synthetic gene circuits, mimicking the way electronic circuits are realized in silico. Basic components were DNA sequences that could be composed, first, into transcription units (TUs) and, then, circuits by exchanging fluxes of molecules, such as PoPS (polymerase per second) and RiPS (ribosomes per seconds) as suggested by Drew Endy. However, it became clear soon that such fluxes were not measurable, which highlighted the limit of using some concepts from electronics to represent biological systems. SBML Level 3 with the comp package permitted us to revise circuit modularity, especially for the modeling of eukaryotic networks. By using the libSBML Python API, TUs-rather than single parts-are encoded in SBML Level 3 files that contain species, reactions, and ports, i.e., the interfaces that permit to wire TUs into circuits. A circuit model consists of a collection of SBML Level 3 files associated with the different TUs plus a "main" file that delineates the circuit structure. Within this framework, there is no more need for any flux of molecules. Here, we present the SBML Level 3-based models and the wet-lab implementations of Boolean NOT gates that make use, in the yeast Saccharomyces cerevisiae, of the bacterial ClpX-ClpP system for protein degradation. This work is the starting point towards a new piece of software for the modular design of eukaryotic gene circuits and shows an alternative way to build genetic Boolean gates.