Tales of Two α-Carboxysomes: the Structure and Assembly of Cargo Rubisco

Carboxysomes are a family of bacterial microcompartments in cyanobacteria and chemoautotrophs. It encapsulates carbonic anhydrase and Ribulose 1,5-bisphosphate carboxylase/oxygenase (Rubisco) catalysing carbon fixation inside a proteinaceous shell. How Rubisco packs into the carboxysomes is unknown. Using cryo-electron tomography and subtomogram averaging, we present 3D organization of Rubisco inside two types of native α-carboxysomes from a marine α-cyanobacterium Cyanobium sp. PCC 7001 and a chemoautotrophic bacterium Halothiobacillus neapolitanus . We determined the structures of Rubiscos within native Halothiobacillus and Cyanobium carboxysomes at 3.3 Å and 3.8 Å resolution respectively and further identified an associated CsoS2 segment. Interestingly, CsoS2 is only associated with a sub-population of Rubiscos that are close to the shell in Halothiobacillus , but with all Rubiscos throughout Cyanobium carboxysome. Moreover, Rubiscos in Cyanobium carboxysomes are organized in three concentric layers whereas Rubiscos in Halothiobacillus carboxysomes are arranged in spiral arrays. Calcium treatment induced a drastic re-organization of Rubiscos, converting these two distinct assemblies into ordered lattice arrays in both α-carboxysomes. Our findings provide critical knowledge of the assembly principles of α-carboxysomes, which may aid in rational design and repurpose of carboxysome structures for new functions. ### Competing Interest Statement The authors have declared no competing interest.