Reprogramming bacterial protein organelles as a nanoreactor for hydrogen production

Abstract

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.

Artificial Organelles: Towards Adding or Restoring Intracellular Activity

“…Such compartments can be experimentally adapted to incorporate new functions or to be functional in higher‐order organisms. Although some engineering into artificial organelles has been done with carboxysomes, [163,164] most of the developments in this field have been made with encapsulins. (Figure 9B).…”

Positioning the Model Bacterial Organelle, the Carboxysome

“…The assembly pathway for a-carboxysomes is less clear, but the fact that a-shell proteins can assemble "ghost" carboxysomes, devoid of core proteins, suggests that inside-out assembly is not obligatory for a-carboxysomes (82,87). Consistently, a recent study engineered a-carboxysome shells (;100-nm diameter) devoid of core proteins (121). These shells were used as nanoreactors to recruit heterologous enzymes for diverse catalytic reactions.…”

Bacterial metabolosomes: new insights into their structure and bioengineering

“…This allows the generation of a CO 2 ‐rich microenvironment within the carboxysome to facilitate Rubisco carboxylation. Our recent studies have provided new insights into the assembly, structure, permeability, physiological regulation and bioengineering of carboxysomes (Sun et al ., 2016; Faulkner et al ., 2017; Fang et al ., 2018; Huang et al ., 2019; Sun et al ., 2019; Faulkner et al ., 2020; Huang et al ., 2020; Li et al ., 2020).…”