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Crystal structures reveal the multi-ligand binding mechanism of Staphylococcus aureus ClfB.

Xiang H, Feng Y, Wang J, Liu B, Chen Y, Liu L, Deng X, Yang M - PLoS Pathog. (2012)

Bottom Line: Structural comparison revealed a conserved glycine-serine-rich (GSR) ClfB binding motif (GSSGXGXXG) within the ligands, which was also found in other human proteins such as Engrailed protein, TCF20 and Dermokine proteins.The results presented here highlight the multi-ligand binding property of ClfB, which is very distinct from other characterized MSCRAMMs to-date.Our results provide a template for the identification of other molecules targeted by S. aureus during its colonization and infection.

View Article: PubMed Central - PubMed

Affiliation: Key Laboratory for Protein Sciences of Ministry of Education, Tsinghua-Peking Center for Life Sciences, School of Life Sciences, Tsinghua University, Beijing, China.

ABSTRACT
Staphylococcus aureus (S. aureus) pathogenesis is a complex process involving a diverse array of extracellular and cell wall components. ClfB, an MSCRAMM (Microbial Surface Components Recognizing Adhesive Matrix Molecules) family surface protein, described as a fibrinogen-binding clumping factor, is a key determinant of S. aureus nasal colonization, but the molecular basis for ClfB-ligand recognition remains unknown. In this study, we solved the crystal structures of apo-ClfB and its complexes with fibrinogen α (Fg α) and cytokeratin 10 (CK10) peptides. Structural comparison revealed a conserved glycine-serine-rich (GSR) ClfB binding motif (GSSGXGXXG) within the ligands, which was also found in other human proteins such as Engrailed protein, TCF20 and Dermokine proteins. Interaction between Dermokine and ClfB was confirmed by subsequent binding assays. The crystal structure of ClfB complexed with a 15-residue peptide derived from Dermokine revealed the same peptide binding mode of ClfB as identified in the crystal structures of ClfB-Fg α and ClfB-CK10. The results presented here highlight the multi-ligand binding property of ClfB, which is very distinct from other characterized MSCRAMMs to-date. The adherence of multiple peptides carrying the GSR motif into the same pocket in ClfB is reminiscent of MHC molecules. Our results provide a template for the identification of other molecules targeted by S. aureus during its colonization and infection. We propose that other MSCRAMMs like ClfA and SdrG also possess multi-ligand binding properties.

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Mechanisms of specifically recognizing repeat 5 of Fg α.A. Superimposition of the Fg α and CK10 peptides. The Fg α and CK10 peptides are shown as sticks, colored in yellow and slate, respectively. Residues highlighted within the boundaries of the red dashed line constitute the segment important for ClfB binding. The consensus amino acids are shown above the peptides. B. Sequence alignment of the repeat 2, 3, 4 and 5 of the Fg α, CK10 (type I cytokeratin 10, residues 473–485 and residues 499–511), K10 (Keratin 10, type I cytoskeletal 10 isoform-1 from Pan troglodytes, residues 501–513), Derm (Dermokine, residues 250–264), TCF20 (TCF20, residues 49–57), EN (Engrailed protein, residues 37–45) and the derived peptide 9. The conserved amino acids are shown in red and the consensus sequence is designated below the sequences. The repeat 2, 3 and 4 of the Fg α which have been proved cannot bind to ClfB are indicated in skyblue.
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ppat-1002751-g005: Mechanisms of specifically recognizing repeat 5 of Fg α.A. Superimposition of the Fg α and CK10 peptides. The Fg α and CK10 peptides are shown as sticks, colored in yellow and slate, respectively. Residues highlighted within the boundaries of the red dashed line constitute the segment important for ClfB binding. The consensus amino acids are shown above the peptides. B. Sequence alignment of the repeat 2, 3, 4 and 5 of the Fg α, CK10 (type I cytokeratin 10, residues 473–485 and residues 499–511), K10 (Keratin 10, type I cytoskeletal 10 isoform-1 from Pan troglodytes, residues 501–513), Derm (Dermokine, residues 250–264), TCF20 (TCF20, residues 49–57), EN (Engrailed protein, residues 37–45) and the derived peptide 9. The conserved amino acids are shown in red and the consensus sequence is designated below the sequences. The repeat 2, 3 and 4 of the Fg α which have been proved cannot bind to ClfB are indicated in skyblue.

Mentions: The Fg α C-terminal domain (amino acids 221–610) of human Fg contains ten 13-residue tandem repeats, within which up to eight residues are glycines or serines [34]. Despite the similar sequences among the repeats, only Fg α5 was shown to be recognized by ClfB [18]. The reason for this was proposed to be the presence of proline or arginine residues in the center of the putative Ω loops in the other repeats though the precise underlying mechanism remains unknown [27]. The crystal structures presented here offer an explanation for this observation. Structural comparison of the two complexes revealed that interactions of the peptides with ClfB are primarily mediated through a conserved motif in the peptides: G-S-S-G-S/T-G-S-X-G (Figure 5A). Sequence alignment of the repeats indicates that Fg α5 differs from the other repeats at the 5th, 7th and 9th positions (Figure 5B). The hydroxyl group of S/T at the 5th position is involved in hydrogen bonding interactions. On the other hand, the size of the residue at this position is limited by its neighboring residues. Thus, other residues except S/T at this position are expected to compromise the interactions between the repeat and ClfB either because of loss of hydrogen bonding interaction or generation of steric hindrance. The 7th position appears to play a role in maintaining the local conformation of the peptide by forming a γ-turn with the 9th position. In the structure of CK10-ClfB complex solved by V.Ganesh et al., the 7th position was replaced with a histidine residue, suggesting that the residue at this position can be varied (Figure S6). The G9 residue was headed to the end of the β-sheet D and the ClfBMet280 and ClfBPro281 in N2 limit residues with any side chain which would generate clash against them. In addition, a turn at the G9 is required to permit the peptide out of the tunnel, explaining why the repeat 2 with an alanine at this position cannot bind to ClfB (Figure 5B) [18].


Crystal structures reveal the multi-ligand binding mechanism of Staphylococcus aureus ClfB.

Xiang H, Feng Y, Wang J, Liu B, Chen Y, Liu L, Deng X, Yang M - PLoS Pathog. (2012)

Mechanisms of specifically recognizing repeat 5 of Fg α.A. Superimposition of the Fg α and CK10 peptides. The Fg α and CK10 peptides are shown as sticks, colored in yellow and slate, respectively. Residues highlighted within the boundaries of the red dashed line constitute the segment important for ClfB binding. The consensus amino acids are shown above the peptides. B. Sequence alignment of the repeat 2, 3, 4 and 5 of the Fg α, CK10 (type I cytokeratin 10, residues 473–485 and residues 499–511), K10 (Keratin 10, type I cytoskeletal 10 isoform-1 from Pan troglodytes, residues 501–513), Derm (Dermokine, residues 250–264), TCF20 (TCF20, residues 49–57), EN (Engrailed protein, residues 37–45) and the derived peptide 9. The conserved amino acids are shown in red and the consensus sequence is designated below the sequences. The repeat 2, 3 and 4 of the Fg α which have been proved cannot bind to ClfB are indicated in skyblue.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3375286&req=5

ppat-1002751-g005: Mechanisms of specifically recognizing repeat 5 of Fg α.A. Superimposition of the Fg α and CK10 peptides. The Fg α and CK10 peptides are shown as sticks, colored in yellow and slate, respectively. Residues highlighted within the boundaries of the red dashed line constitute the segment important for ClfB binding. The consensus amino acids are shown above the peptides. B. Sequence alignment of the repeat 2, 3, 4 and 5 of the Fg α, CK10 (type I cytokeratin 10, residues 473–485 and residues 499–511), K10 (Keratin 10, type I cytoskeletal 10 isoform-1 from Pan troglodytes, residues 501–513), Derm (Dermokine, residues 250–264), TCF20 (TCF20, residues 49–57), EN (Engrailed protein, residues 37–45) and the derived peptide 9. The conserved amino acids are shown in red and the consensus sequence is designated below the sequences. The repeat 2, 3 and 4 of the Fg α which have been proved cannot bind to ClfB are indicated in skyblue.
Mentions: The Fg α C-terminal domain (amino acids 221–610) of human Fg contains ten 13-residue tandem repeats, within which up to eight residues are glycines or serines [34]. Despite the similar sequences among the repeats, only Fg α5 was shown to be recognized by ClfB [18]. The reason for this was proposed to be the presence of proline or arginine residues in the center of the putative Ω loops in the other repeats though the precise underlying mechanism remains unknown [27]. The crystal structures presented here offer an explanation for this observation. Structural comparison of the two complexes revealed that interactions of the peptides with ClfB are primarily mediated through a conserved motif in the peptides: G-S-S-G-S/T-G-S-X-G (Figure 5A). Sequence alignment of the repeats indicates that Fg α5 differs from the other repeats at the 5th, 7th and 9th positions (Figure 5B). The hydroxyl group of S/T at the 5th position is involved in hydrogen bonding interactions. On the other hand, the size of the residue at this position is limited by its neighboring residues. Thus, other residues except S/T at this position are expected to compromise the interactions between the repeat and ClfB either because of loss of hydrogen bonding interaction or generation of steric hindrance. The 7th position appears to play a role in maintaining the local conformation of the peptide by forming a γ-turn with the 9th position. In the structure of CK10-ClfB complex solved by V.Ganesh et al., the 7th position was replaced with a histidine residue, suggesting that the residue at this position can be varied (Figure S6). The G9 residue was headed to the end of the β-sheet D and the ClfBMet280 and ClfBPro281 in N2 limit residues with any side chain which would generate clash against them. In addition, a turn at the G9 is required to permit the peptide out of the tunnel, explaining why the repeat 2 with an alanine at this position cannot bind to ClfB (Figure 5B) [18].

Bottom Line: Structural comparison revealed a conserved glycine-serine-rich (GSR) ClfB binding motif (GSSGXGXXG) within the ligands, which was also found in other human proteins such as Engrailed protein, TCF20 and Dermokine proteins.The results presented here highlight the multi-ligand binding property of ClfB, which is very distinct from other characterized MSCRAMMs to-date.Our results provide a template for the identification of other molecules targeted by S. aureus during its colonization and infection.

View Article: PubMed Central - PubMed

Affiliation: Key Laboratory for Protein Sciences of Ministry of Education, Tsinghua-Peking Center for Life Sciences, School of Life Sciences, Tsinghua University, Beijing, China.

ABSTRACT
Staphylococcus aureus (S. aureus) pathogenesis is a complex process involving a diverse array of extracellular and cell wall components. ClfB, an MSCRAMM (Microbial Surface Components Recognizing Adhesive Matrix Molecules) family surface protein, described as a fibrinogen-binding clumping factor, is a key determinant of S. aureus nasal colonization, but the molecular basis for ClfB-ligand recognition remains unknown. In this study, we solved the crystal structures of apo-ClfB and its complexes with fibrinogen α (Fg α) and cytokeratin 10 (CK10) peptides. Structural comparison revealed a conserved glycine-serine-rich (GSR) ClfB binding motif (GSSGXGXXG) within the ligands, which was also found in other human proteins such as Engrailed protein, TCF20 and Dermokine proteins. Interaction between Dermokine and ClfB was confirmed by subsequent binding assays. The crystal structure of ClfB complexed with a 15-residue peptide derived from Dermokine revealed the same peptide binding mode of ClfB as identified in the crystal structures of ClfB-Fg α and ClfB-CK10. The results presented here highlight the multi-ligand binding property of ClfB, which is very distinct from other characterized MSCRAMMs to-date. The adherence of multiple peptides carrying the GSR motif into the same pocket in ClfB is reminiscent of MHC molecules. Our results provide a template for the identification of other molecules targeted by S. aureus during its colonization and infection. We propose that other MSCRAMMs like ClfA and SdrG also possess multi-ligand binding properties.

Show MeSH
Related in: MedlinePlus