Abstract
Bacterial microcompartments (BMCs) are promising natural protein structures for applications that require the segregation of certain metabolic functions or molecular species in a defined microenvironment. To understand how endogenous cargos are packaged inside the protein shell is key for using BMCs as nano-scale reactors or delivery vesicles. In this report, we studied the encapsulation of RuBisCO into the α-type carboxysome from Halothiobacillus neapolitan. Our experimental data revealed that the CsoS2 scaffold proteins engage RuBisCO enzyme through an interaction with the small subunit (CbbS). In addition, the N domain of the large subunit (CbbL) of RuBisCO interacts with all shell proteins that can form the hexamers. The binding affinity between the N domain of CbbL and one of the major shell proteins, CsoS1C, is within the submicromolar range. The absence of the N domain also prevented the encapsulation of the rest of the RuBisCO subunits. Our findings complete the picture of how RuBisCOs are encapsulated into the α-type carboxysome and provide insights for future studies and engineering of carboxysome as a protein shell.
Highlights
Bacterial microcompartments (BMCs) are natural proteinaceous cell organelles that encapsulate functionally relevant enzymes to execute designated steps of biological pathways[1,2,3,4]
Mechanistic studies on protein encapsulation into the Pdu and Eut BMCs of S. enterica revealed that the N-termini of functional proteins, such as PduA and EutG, serve as signal peptides to interact with shell proteins and are responsible for the formation of BMCs around the cargos[18,22,23]
Bands of minor shell proteins (CsoS1D, CsoS4A, and CsoS4B) and encapsulated carbonic anhydrase (CsoS3) could not be unambiguously identified, which is consistent with a previous report[32]
Summary
Bacterial microcompartments (BMCs) are natural proteinaceous cell organelles that encapsulate functionally relevant enzymes to execute designated steps of biological pathways[1,2,3,4]. Mechanistic studies on protein encapsulation into the Pdu and Eut BMCs of S. enterica revealed that the N-termini of functional proteins, such as PduA and EutG, serve as signal peptides to interact with shell proteins and are responsible for the formation of BMCs around the cargos[18,22,23]. A similar mechanism was uncovered in metabolosomes from other bacterial species, albeit the signal peptide may reside on the C-termini[19,24] This knowledge guided efforts toward the engineering of artificial interaction motifs for foreign cargo protein www.nature.com/scientificreports/. The objective of the present work is to understand the encapsulation mechanism of α-type carboxysome through the study of interactions between BMC shell/scaffold proteins and cargo proteins.
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