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.


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Research sample  Fig. 10 are provided as a Source Data file. All data are available from the corresponding author upon request. Uniprot database was used to search the unique of the target peptides of each carboxysomal protein.
The sample size for TEM data were determined by the number of available images which were of high quality. The sample size for SDS-PAGE, Western blot, Mass spectrometry, Gas chromatography, Dynamic light scattering, DO analysis was not predetermined using statistical methods. At least three biological replicates were analyzed by Immunoblot, Dynamic light scattering and Gas chromatography and DO measurements. Similar results were obtained from the replicates of each experimental analysis.
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