| Affiliation: | 1. Department of Cell Biology and Neuroscience & Institute for Quantitative Biomedicine, Rutgers University, Piscataway, NJ 08854, USA;2. Verna and Marrs Mclean Department of Biochemistry and Molecular Biology, Baylor College of Medicine, Houston, TX 77030, USA;3. Department of Pharmacology and Toxicology, University of Texas Medical Branch, Galveston, TX 77555, USA;4. Department of Biochemistry and Molecular Biology, University of Texas Medical Branch, Galveston, TX 77555, USA;5. Sealy Center for Structural Biology and Molecular Biophysics, University of Texas Medical Branch, Galveston, TX 77555, USA;6. Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02139, USA;7. Division of CryoEM and Bioimaging, SSRL, SLAC National Accelerator Laboratory, Stanford University, Menlo Park, CA 94025, USA;8. Departments of Bioengineering and of Microbiology and Immunoplogy, James H. Clark Center, Stanford University, Stanford, CA 94305, USA |
| Abstract: | Cyanobacteria are photosynthetic organisms responsible for ~ 25% of the organic carbon fixation on earth. A key step in carbon fixation is catalyzed by ribulose bisphosphate carboxylase/oxygenase (RuBisCO), the most abundant enzyme in the biosphere. Applying Zernike phase-contrast electron cryo-tomography and automated annotation, we identified individual RuBisCO molecules and their assembly intermediates leading to the formation of carboxysomes inside Syn5 cyanophage infected cyanobacteria Synechococcus sp. WH8109 cells. Surprisingly, more RuBisCO molecules were found to be present as cytosolic free-standing complexes or clusters than as packaged assemblies inside carboxysomes. Cytosolic RuBisCO clusters and partially assembled carboxysomes identified in the cell tomograms support a concurrent assembly model involving both the protein shell and the enclosed RuBisCO. In mature carboxysomes, RuBisCO is neither randomly nor strictly icosahedrally packed within protein shells of variable sizes. A time-averaged molecular dynamics simulation showed a semi-liquid probability distribution of the RuBisCO in carboxysomes and correlated well with carboxysome subtomogram averages. Our structural observations reveal the various stages of RuBisCO assemblies, which could be important for understanding cellular function. |
