Bottom-up Construction of a Primordial Carboxysome Mimic.

We have constructed a synthetic mimic of the carboxysome, a cyanobacterial carbon-fixing organelle. Using an electrostatic tagging system, we coencapsulated the two key carboxysomal enzymes, ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO) and carbonic anhydrase (CA), in an engineered protein cage based on lumazine synthase. A statistically significant kinetic effect of coencapsulated CA on RuBisCO activity was not observed under ambient or oxygen saturated conditions, suggesting that enzyme proximity alone may not be the key determinant in carboxysome function. The capsid shell protected the enzyme from proteolytic damage, a factor that could have provided early cyanobacteria with an evolutionary benefit. Our strategy to coencapsulate different proteins can easily be extended to other sequentially acting enzymes and lays down principles for developing artificial organelles to control biosynthetic pathways in vivo.