Reprogramming encapsulins into modular carbon-fixing nanocompartments

Introducing CO2-concentrating mechanisms (CCM) into C3 crops represents a major frontier in synthetic biology with potential to enhance photosynthetic efficiency and yields. Despite decades of progress in elucidating CCM components, mechanisms and genetics (including structures of native Rubisco-containing compartments), installing algal pyrenoids or cyanobacterial carboxysomes into plants remains a formidable challenge. This is due to the requirement for chloroplast engineering to facilitate sufficient expression, and specificity of condensate proteins that impedes use of heterologous Rubiscos without extensive genetic redesign. Here, we present a modular streamlined alternative, a synthetic system using encapsulin nanocompartments from Quasibacillus thermotolerans (QtEnc). By fusing a short cargo-loading peptide to diverse Rubisco isoforms, we achieve targeted encapsulation within QtEnc while retaining CO2-fixing activity. Our isoform-agnostic design establishes a foundation for constructing plant-compatible synthetic carboxysome mimics. While carbonic anhydrase remains to be incorporated, our system offers a simpler tractable path towards integrating a functional CCM in crops. Installing algal pyrenoids or cyanobacterial carboxysomes into plants to enhance photosynthetic efficiency remains a formidable challenge. Here, the authors report the construction of CO2-fixing nanocompartments by encapsulating Rubisco using encapsulin from Quasibacillus thermotolerans.