Reprogramming bacterial protein organelles as a nanoreactor for hydrogen production

Compartmentalization is a ubiquitous building principle in cells, which permits segregation of biological elements and reactions. The carboxysome is a specialized bacterial organelle that encapsulates enzymes into a virus-like protein shell and plays essential roles in photosynthetic carbon fixation. The naturally designed architecture, semi-permeability, and catalytic improvement of carboxysomes have inspired rational design and engineering of new nanomaterials to incorporate desired enzymes into the protein shell for enhanced catalytic performance. Here, we build large, intact carboxysome shells (over 90 nm in diameter) in the industrial microorganism Escherichia coli by expressing a set of carboxysome protein-encoding genes. We develop strategies for enzyme activation, shell self-assembly, and cargo encapsulation to construct a robust nanoreactor that incorporates catalytically active [FeFe]-hydrogenases and functional partners within the empty shell for the production of hydrogen. We show that shell encapsulation and the internal microenvironment of the new catalyst facilitate hydrogen production of the encapsulated oxygen-sensitive hydrogenases. The study provides insights into the assembly and formation of carboxysomes and paves the way for engineering carboxysome shell-based nanoreactors to recruit specific enzymes for diverse catalytic reactions.