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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.


Folding predictions for MIT9313 CsoS2. (a) Fold-Index prediction; (b) PONDR prediction; (c) ribbon presentation of ab initio folding prediction by QUARK for each M-repeat, shown in a rainbow spectrum from N-terminus (blue) to C-terminus (red).
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life-05-01141-f008: Folding predictions for MIT9313 CsoS2. (a) Fold-Index prediction; (b) PONDR prediction; (c) ribbon presentation of ab initio folding prediction by QUARK for each M-repeat, shown in a rainbow spectrum from N-terminus (blue) to C-terminus (red).

Mentions: Protein Basic Local Alignment Search Tool (BLASTp) searches using the primary structures of Hnea, MED4 or MIT9313 CsoS2 against Protein Data Bank (PDB) with an E-value cut-off of 1.0 returned no hits. Therefore, we undertook in silico tertiary structure prediction for all available CsoS2 sequences. The predictions resulting from two different algorithms are similar and the majority of sampled CsoS2 orthologs are predicted to be disordered. Only a few exceptions have more than one region (>50 residues) predicted to adopt a local fold (CsoS2 from Acidimicrobium ferrooxidans DSM 10331, Bradyrhizobium sp. BTAi1 and Thermithiobacillus tepidarius DSM 3134). Predictions for the MIT9313 CsoS2 are shown in Figure 8a,b as examples. Using FoldIndex, more than 70% of the sequence is predicted as unfolded (in red) with three locally folded regions (in green) (Figure 8a), each of which roughly corresponds to one of the three big dips in the PONDR prediction (Figure 8b), indicating potential order locally or order formed when a binding partner is present. The first is located in the first M-repeat; the second occurs between the 5th and 6th M-repeats, and the last is at the beginning of the C-region. The consistency in prediction of locally-ordered regions suggests the possibility of the M-region adopting a beads-on-a-string conformation, with beads representing local units of tertiary structure. We performed ab initio protein structure prediction for each M-repeat with flanking sequences. Excluding the sixth repeat, four out of five long M-repeats all adopt a [(β-strand)4-6-(α-helix)1-2] conformation (Figure 8c). The third M-repeat (short form) also adopts a (β-strand)4-6 conformation but lacks the α-helix portion due to the absence of 10 amino acid C-terminal extension (Figure 6d). Together, these results suggest that CsoS2 is highly flexible, and a beads-on-a-string conformation is possible at least for the M-region.


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)

Folding predictions for MIT9313 CsoS2. (a) Fold-Index prediction; (b) PONDR prediction; (c) ribbon presentation of ab initio folding prediction by QUARK for each M-repeat, shown in a rainbow spectrum from N-terminus (blue) to C-terminus (red).
© Copyright Policy
Related In: Results  -  Collection

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

life-05-01141-f008: Folding predictions for MIT9313 CsoS2. (a) Fold-Index prediction; (b) PONDR prediction; (c) ribbon presentation of ab initio folding prediction by QUARK for each M-repeat, shown in a rainbow spectrum from N-terminus (blue) to C-terminus (red).
Mentions: Protein Basic Local Alignment Search Tool (BLASTp) searches using the primary structures of Hnea, MED4 or MIT9313 CsoS2 against Protein Data Bank (PDB) with an E-value cut-off of 1.0 returned no hits. Therefore, we undertook in silico tertiary structure prediction for all available CsoS2 sequences. The predictions resulting from two different algorithms are similar and the majority of sampled CsoS2 orthologs are predicted to be disordered. Only a few exceptions have more than one region (>50 residues) predicted to adopt a local fold (CsoS2 from Acidimicrobium ferrooxidans DSM 10331, Bradyrhizobium sp. BTAi1 and Thermithiobacillus tepidarius DSM 3134). Predictions for the MIT9313 CsoS2 are shown in Figure 8a,b as examples. Using FoldIndex, more than 70% of the sequence is predicted as unfolded (in red) with three locally folded regions (in green) (Figure 8a), each of which roughly corresponds to one of the three big dips in the PONDR prediction (Figure 8b), indicating potential order locally or order formed when a binding partner is present. The first is located in the first M-repeat; the second occurs between the 5th and 6th M-repeats, and the last is at the beginning of the C-region. The consistency in prediction of locally-ordered regions suggests the possibility of the M-region adopting a beads-on-a-string conformation, with beads representing local units of tertiary structure. We performed ab initio protein structure prediction for each M-repeat with flanking sequences. Excluding the sixth repeat, four out of five long M-repeats all adopt a [(β-strand)4-6-(α-helix)1-2] conformation (Figure 8c). The third M-repeat (short form) also adopts a (β-strand)4-6 conformation but lacks the α-helix portion due to the absence of 10 amino acid C-terminal extension (Figure 6d). Together, these results suggest that CsoS2 is highly flexible, and a beads-on-a-string conformation is possible at least for the M-region.

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.