Quantitative Measurement of Molecular Permeability to a Synthetic Bacterial Microcompartment Shell System

Naturally evolved and synthetically designed forms of compartmentalization benefit encapsulated function by increasing local concentrations of substrates and protecting cargo from destabilizing environments and inhibitors. Crucial to understanding the fundamental principles of compartmentalization are experimental systems enabling the measurement of the permeability rates of small molecules. Here, we report the experimental measurement of the small-molecule permeability of a 40 nm icosahedral bacterial microcompartment shell. This was accomplished by heterologous loading of light-producing luciferase enzymes and kinetic measurement of luminescence using stopped-flow spectrophotometry. Compared to free enzyme, the luminescence signal kinetics was slower when the luciferase was encapsulated in bacterial microcompartment shells. The results indicate that substrates and products can still exchange across the shell, and modeling of the experimental data suggest that a 50× permeability rate increase occurs when shell vertices were vacant. Overall, our results suggest design considerations for the construction of heterologous bacterial microcompartment shell systems and compartmentalized function at the nanoscale.