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Advances in Understanding Carboxysome Assembly in Prochlorococcus and Synechococcus Implicate CsoS2 as a Critical Component.

Cai F, Dou Z, Bernstein SL, Leverenz R, Williams EB, Heinhorst S, Shively J, Cannon GC, Kerfeld CA - Life (Basel) (2015)

Bottom Line: Two types of carboxysome, α and β, encapsulating form IA and form IB d-ribulose-1,5-bisphosphate carboxylase/oxygenase, respectively, differ in gene organization and associated proteins.Based on our results from bioinformatic, biophysical, genetic and biochemical approaches, including peptide array scanning for protein-protein interactions, we propose a model for CsoS2 function and its spatial location in the α-carboxysome.Analogies between the pathway for β-carboxysome biogenesis and our model for α-carboxysome assembly are discussed.

View Article: PubMed Central - PubMed

Affiliation: Department of Plant and Microbial Biology, University of California, Berkeley, CA 94720, USA. fcai@lbl.gov.

ABSTRACT
The marine Synechococcus and Prochlorococcus are the numerically dominant cyanobacteria in the ocean and important in global carbon fixation. They have evolved a CO2-concentrating-mechanism, of which the central component is the carboxysome, a self-assembling proteinaceous organelle. Two types of carboxysome, α and β, encapsulating form IA and form IB d-ribulose-1,5-bisphosphate carboxylase/oxygenase, respectively, differ in gene organization and associated proteins. In contrast to the β-carboxysome, the assembly process of the α-carboxysome is enigmatic. Moreover, an absolutely conserved α-carboxysome protein, CsoS2, is of unknown function and has proven recalcitrant to crystallization. Here, we present studies on the CsoS2 protein in three model organisms and show that CsoS2 is vital for α-carboxysome biogenesis. The primary structure of CsoS2 appears tripartite, composed of an N-terminal, middle (M)-, and C-terminal region. Repetitive motifs can be identified in the N- and M-regions. Multiple lines of evidence suggest CsoS2 is highly flexible, possibly an intrinsically disordered protein. Based on our results from bioinformatic, biophysical, genetic and biochemical approaches, including peptide array scanning for protein-protein interactions, we propose a model for CsoS2 function and its spatial location in the α-carboxysome. Analogies between the pathway for β-carboxysome biogenesis and our model for α-carboxysome assembly are discussed.

No MeSH data available.


Native agarose gel electrophoresis of Hnea recombinant carboxysome protein and Hnea RuBisCO mixtures. Lanes from top to bottom: rCsoS2 (20 µL at 0.6 mg/mL), RuBisCO (20 µL at 1.1 mg/mL), rCsoS1A (20 µL at 0.7 mg/mL), BSA (20 µL at 1.0 mg/mL), rCsoS2 w. RuBisCO and rCsoS1A (20 µL each), rCsoS2 w. RuBisCO, rCsoS1A and BSA (20 µL each), and rCsoS2 w. BSA and rCsoS1A (20 µL each). By itself, the positively charged rCsoS2 migrates to the negative electrode; rCsoS1A and RuBisCO migrate to the positive electrode. When mixed, rCsoS2 drags its interaction partners, but not BSA, towards the negative electrode.
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life-05-01141-f010: Native agarose gel electrophoresis of Hnea recombinant carboxysome protein and Hnea RuBisCO mixtures. Lanes from top to bottom: rCsoS2 (20 µL at 0.6 mg/mL), RuBisCO (20 µL at 1.1 mg/mL), rCsoS1A (20 µL at 0.7 mg/mL), BSA (20 µL at 1.0 mg/mL), rCsoS2 w. RuBisCO and rCsoS1A (20 µL each), rCsoS2 w. RuBisCO, rCsoS1A and BSA (20 µL each), and rCsoS2 w. BSA and rCsoS1A (20 µL each). By itself, the positively charged rCsoS2 migrates to the negative electrode; rCsoS1A and RuBisCO migrate to the positive electrode. When mixed, rCsoS2 drags its interaction partners, but not BSA, towards the negative electrode.

Mentions: Similarly, protein-protein interactions were observed upon mixing of soluble recombinant Hnea CsoS2, the shell protein CsoS1A, and Hnea RuBisCO purified from the native source. In native agarose gels, the positively charged rCsoS2 migrates to the negative electrode; rCsoS1A and RuBisCO migrate to positive electrode (Figure 10). When mixed, all of the proteins, presumably as a complex, migrate to the negative electrode. Commercially available BSA was used as a control and it only migrates to the positive electrode in the absence or presence of CsoS2 in native agarose gels (Figure 10), which further supports the hypothesis that protein-protein interactions between CsoS2 and shell proteins or RuBisCO are specific.


Advances in Understanding Carboxysome Assembly in Prochlorococcus and Synechococcus Implicate CsoS2 as a Critical Component.

Cai F, Dou Z, Bernstein SL, Leverenz R, Williams EB, Heinhorst S, Shively J, Cannon GC, Kerfeld CA - Life (Basel) (2015)

Native agarose gel electrophoresis of Hnea recombinant carboxysome protein and Hnea RuBisCO mixtures. Lanes from top to bottom: rCsoS2 (20 µL at 0.6 mg/mL), RuBisCO (20 µL at 1.1 mg/mL), rCsoS1A (20 µL at 0.7 mg/mL), BSA (20 µL at 1.0 mg/mL), rCsoS2 w. RuBisCO and rCsoS1A (20 µL each), rCsoS2 w. RuBisCO, rCsoS1A and BSA (20 µL each), and rCsoS2 w. BSA and rCsoS1A (20 µL each). By itself, the positively charged rCsoS2 migrates to the negative electrode; rCsoS1A and RuBisCO migrate to the positive electrode. When mixed, rCsoS2 drags its interaction partners, but not BSA, towards the negative electrode.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4499774&req=5

life-05-01141-f010: Native agarose gel electrophoresis of Hnea recombinant carboxysome protein and Hnea RuBisCO mixtures. Lanes from top to bottom: rCsoS2 (20 µL at 0.6 mg/mL), RuBisCO (20 µL at 1.1 mg/mL), rCsoS1A (20 µL at 0.7 mg/mL), BSA (20 µL at 1.0 mg/mL), rCsoS2 w. RuBisCO and rCsoS1A (20 µL each), rCsoS2 w. RuBisCO, rCsoS1A and BSA (20 µL each), and rCsoS2 w. BSA and rCsoS1A (20 µL each). By itself, the positively charged rCsoS2 migrates to the negative electrode; rCsoS1A and RuBisCO migrate to the positive electrode. When mixed, rCsoS2 drags its interaction partners, but not BSA, towards the negative electrode.
Mentions: Similarly, protein-protein interactions were observed upon mixing of soluble recombinant Hnea CsoS2, the shell protein CsoS1A, and Hnea RuBisCO purified from the native source. In native agarose gels, the positively charged rCsoS2 migrates to the negative electrode; rCsoS1A and RuBisCO migrate to positive electrode (Figure 10). When mixed, all of the proteins, presumably as a complex, migrate to the negative electrode. Commercially available BSA was used as a control and it only migrates to the positive electrode in the absence or presence of CsoS2 in native agarose gels (Figure 10), which further supports the hypothesis that protein-protein interactions between CsoS2 and shell proteins or RuBisCO are specific.

Bottom Line: Two types of carboxysome, α and β, encapsulating form IA and form IB d-ribulose-1,5-bisphosphate carboxylase/oxygenase, respectively, differ in gene organization and associated proteins.Based on our results from bioinformatic, biophysical, genetic and biochemical approaches, including peptide array scanning for protein-protein interactions, we propose a model for CsoS2 function and its spatial location in the α-carboxysome.Analogies between the pathway for β-carboxysome biogenesis and our model for α-carboxysome assembly are discussed.

View Article: PubMed Central - PubMed

Affiliation: Department of Plant and Microbial Biology, University of California, Berkeley, CA 94720, USA. fcai@lbl.gov.

ABSTRACT
The marine Synechococcus and Prochlorococcus are the numerically dominant cyanobacteria in the ocean and important in global carbon fixation. They have evolved a CO2-concentrating-mechanism, of which the central component is the carboxysome, a self-assembling proteinaceous organelle. Two types of carboxysome, α and β, encapsulating form IA and form IB d-ribulose-1,5-bisphosphate carboxylase/oxygenase, respectively, differ in gene organization and associated proteins. In contrast to the β-carboxysome, the assembly process of the α-carboxysome is enigmatic. Moreover, an absolutely conserved α-carboxysome protein, CsoS2, is of unknown function and has proven recalcitrant to crystallization. Here, we present studies on the CsoS2 protein in three model organisms and show that CsoS2 is vital for α-carboxysome biogenesis. The primary structure of CsoS2 appears tripartite, composed of an N-terminal, middle (M)-, and C-terminal region. Repetitive motifs can be identified in the N- and M-regions. Multiple lines of evidence suggest CsoS2 is highly flexible, possibly an intrinsically disordered protein. Based on our results from bioinformatic, biophysical, genetic and biochemical approaches, including peptide array scanning for protein-protein interactions, we propose a model for CsoS2 function and its spatial location in the α-carboxysome. Analogies between the pathway for β-carboxysome biogenesis and our model for α-carboxysome assembly are discussed.

No MeSH data available.