Design Principles for a Compartmentalized Enzyme Cascade Reaction

Compartmentalization of enzyme cascade reactions can both create a safe space for volatile reaction intermediates, where they are protected from cellular degradation mechanisms, and a quarantine for toxic intermediates, where they are prevented from impeding cellular function. The quantitative understanding of biological compartments and the design of bioengineered compartments and synthetic cells would be facilitated by a general and concise model. Most existing models for studying compartmentalized cascades focus on a specific biological system and are highly detailed. In this study, we develop a simple model for a compartmentalized two-enzyme cascade reaction and analyze it in the well-mixed, steady-state regime. An immediate and intuitive result is that the fundamental parameter governing compartment cascade throughput is the resistance to diffusion of substrate and intermediate molecules across the compartment boundary. We then use this model to develop a design process for a compartmentalized cascade, where intermediate loss is minimized while maintaining a desired product outflux. Finally, the model also reveals that there is a critical threshold at which compartmentalization provides benefits over the free-solution reaction. Our model not only provides many insights into the design of compartmentalized cascade reactions, but also captures the essential physics of the problem, as it can replicate the results of more complex models.