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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.


A Hidden Markov model (HMM) logo for all α-Cyanobacterial CsoS2 orthologs. The Y-axis represents the information content (aka relative entropy), and the letters divide the stack height according to their estimated probability at a given position. MED4 and MIT9313 CsoS2 sequences are aligned to the corresponding position on the logo, and the predicted secondary structural motifs are colored red and orange for α-helices and β-strands, respectively. N- and M-region repeats are indicated by cyan and green underlining, respectively. Short repeats (3 amino acids) that occur three units per group (except in the last group) are outlined in light-gray boxes. Relatively conserved residues of the C-region are underlined in red. Putative transition areas between three regions are indicated by brown arrows. For demonstration purposes only, a simplified presentation of results from the protein-binding assay against MIT9313 CsoS2 peptide array (see Section 2.12) are mapped onto the logo. The starting position of peptides among all positive hits is marked with RuBisCO, CsoS1 or CsoS1D symbols only if the averaged signal intensity (1) ranks in the top 10 out of all positive hits or (2) is a local maximum with >5 sequential positive hits. The saturation of each symbol is relative to its fraction ratio to the maximum signal intensity (as 100% saturation) of all positive hits from a given binding assay.
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life-05-01141-f005: A Hidden Markov model (HMM) logo for all α-Cyanobacterial CsoS2 orthologs. The Y-axis represents the information content (aka relative entropy), and the letters divide the stack height according to their estimated probability at a given position. MED4 and MIT9313 CsoS2 sequences are aligned to the corresponding position on the logo, and the predicted secondary structural motifs are colored red and orange for α-helices and β-strands, respectively. N- and M-region repeats are indicated by cyan and green underlining, respectively. Short repeats (3 amino acids) that occur three units per group (except in the last group) are outlined in light-gray boxes. Relatively conserved residues of the C-region are underlined in red. Putative transition areas between three regions are indicated by brown arrows. For demonstration purposes only, a simplified presentation of results from the protein-binding assay against MIT9313 CsoS2 peptide array (see Section 2.12) are mapped onto the logo. The starting position of peptides among all positive hits is marked with RuBisCO, CsoS1 or CsoS1D symbols only if the averaged signal intensity (1) ranks in the top 10 out of all positive hits or (2) is a local maximum with >5 sequential positive hits. The saturation of each symbol is relative to its fraction ratio to the maximum signal intensity (as 100% saturation) of all positive hits from a given binding assay.

Mentions: Although CsoS2 has the least conserved primary structure among all α-carboxysome proteins [27,28], some unusual sequence features are shared among Hnea CsoS2 and its counterparts in Prochlorococcus strains. First, the three CsoS2 proteins from Hnea, MED4 and MIT9313 have unusually high pI values (Table 1). Secondary structure predictions suggest that CsoS2 can be divided into three regions (Figure 5): an approximately 250 amino acid N-terminal region predicted to be enriched in α-helices and having an even higher pI than the full-length proteins (≥ 1 unit difference); an over 350 amino acid middle (M) region predicted to be predominantly composed of β-strands and having a slightly basic pI (7–8); and a (~170 amino acid) C-terminal region. The pI values of the C-terminal region for MED4, MIT9313 and Hnea CsoS2 are 5.26, 7.05, and 9.56, respectively.


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)

A Hidden Markov model (HMM) logo for all α-Cyanobacterial CsoS2 orthologs. The Y-axis represents the information content (aka relative entropy), and the letters divide the stack height according to their estimated probability at a given position. MED4 and MIT9313 CsoS2 sequences are aligned to the corresponding position on the logo, and the predicted secondary structural motifs are colored red and orange for α-helices and β-strands, respectively. N- and M-region repeats are indicated by cyan and green underlining, respectively. Short repeats (3 amino acids) that occur three units per group (except in the last group) are outlined in light-gray boxes. Relatively conserved residues of the C-region are underlined in red. Putative transition areas between three regions are indicated by brown arrows. For demonstration purposes only, a simplified presentation of results from the protein-binding assay against MIT9313 CsoS2 peptide array (see Section 2.12) are mapped onto the logo. The starting position of peptides among all positive hits is marked with RuBisCO, CsoS1 or CsoS1D symbols only if the averaged signal intensity (1) ranks in the top 10 out of all positive hits or (2) is a local maximum with >5 sequential positive hits. The saturation of each symbol is relative to its fraction ratio to the maximum signal intensity (as 100% saturation) of all positive hits from a given binding assay.
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Related In: Results  -  Collection

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

life-05-01141-f005: A Hidden Markov model (HMM) logo for all α-Cyanobacterial CsoS2 orthologs. The Y-axis represents the information content (aka relative entropy), and the letters divide the stack height according to their estimated probability at a given position. MED4 and MIT9313 CsoS2 sequences are aligned to the corresponding position on the logo, and the predicted secondary structural motifs are colored red and orange for α-helices and β-strands, respectively. N- and M-region repeats are indicated by cyan and green underlining, respectively. Short repeats (3 amino acids) that occur three units per group (except in the last group) are outlined in light-gray boxes. Relatively conserved residues of the C-region are underlined in red. Putative transition areas between three regions are indicated by brown arrows. For demonstration purposes only, a simplified presentation of results from the protein-binding assay against MIT9313 CsoS2 peptide array (see Section 2.12) are mapped onto the logo. The starting position of peptides among all positive hits is marked with RuBisCO, CsoS1 or CsoS1D symbols only if the averaged signal intensity (1) ranks in the top 10 out of all positive hits or (2) is a local maximum with >5 sequential positive hits. The saturation of each symbol is relative to its fraction ratio to the maximum signal intensity (as 100% saturation) of all positive hits from a given binding assay.
Mentions: Although CsoS2 has the least conserved primary structure among all α-carboxysome proteins [27,28], some unusual sequence features are shared among Hnea CsoS2 and its counterparts in Prochlorococcus strains. First, the three CsoS2 proteins from Hnea, MED4 and MIT9313 have unusually high pI values (Table 1). Secondary structure predictions suggest that CsoS2 can be divided into three regions (Figure 5): an approximately 250 amino acid N-terminal region predicted to be enriched in α-helices and having an even higher pI than the full-length proteins (≥ 1 unit difference); an over 350 amino acid middle (M) region predicted to be predominantly composed of β-strands and having a slightly basic pI (7–8); and a (~170 amino acid) C-terminal region. The pI values of the C-terminal region for MED4, MIT9313 and Hnea CsoS2 are 5.26, 7.05, and 9.56, respectively.

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.