Conserved and repetitive motifs in an intrinsically disordered protein drive α-carboxysome assembly

Abstract

AbstractAll photosynthetic bacteria and some chemoautotrophic bacteria fix CO2into sugars in specialized proteinaceous compartments called carboxysomes. Carboxysomes enclose the enzymes Rubisco and carbonic anhydrase inside a layer of shell proteins to increase the CO2concentration for efficient carbon fixation by Rubisco. In the α-carboxysome lineage, a disordered and highly repetitive protein named CsoS2 is essential for carboxysome formation and function. Without it, the bacteria are unable to fix enough carbon to grow in air. How a protein lacking structure serves as the architectural scaffold for such a vital cellular compartment remains unknown. In this study, we identify key residues in CsoS2 that are necessary for building functional α-carboxysomesin vivo. These highly conserved and repetitive residues, VTG and Y, contribute to the interaction between CsoS2 and shell proteins. We also demonstratein vitroreconstitution of the α-carboxysome into spherical condensates with CsoS2, Rubisco, and shell proteins, and show the utility of reconstitution as a biochemical tool to study carboxysome biogenesis. The precise self-assembly of thousands of proteins is crucial for carboxysome formation, and understanding this process could enable their use in alternative biological hosts or industrial processes as effective tools to fix carbon.