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Structural analysis of the interactions between paxillin LD motifs and alpha-parvin.

Lorenz S, Vakonakis I, Lowe ED, Campbell ID, Noble ME, Hoellerer MK - Structure (2008)

Bottom Line: Cocrystal structures with these LD motifs reveal the molecular details of their interactions with a common binding site on alpha-parvin-CH(C), which is located at the rim of the canonical fold and includes part of the inter-CH domain linker.Surprisingly, this binding site can accommodate LD motifs in two antiparallel orientations.Taken together, these results reveal an unusual degree of binding degeneracy in the paxillin/alpha-parvin system that may facilitate the assembly of dynamic signaling complexes in the cell.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Molecular Biophysics, University of Oxford, Oxford OX1 3QU, United Kingdom.

ABSTRACT
The adaptor protein paxillin contains five conserved leucine-rich (LD) motifs that interact with a variety of focal adhesion proteins, such as alpha-parvin. Here, we report the first crystal structure of the C-terminal calponin homology domain (CH(C)) of alpha-parvin at 1.05 A resolution and show that it is able to bind all the LD motifs, with some selectivity for LD1, LD2, and LD4. Cocrystal structures with these LD motifs reveal the molecular details of their interactions with a common binding site on alpha-parvin-CH(C), which is located at the rim of the canonical fold and includes part of the inter-CH domain linker. Surprisingly, this binding site can accommodate LD motifs in two antiparallel orientations. Taken together, these results reveal an unusual degree of binding degeneracy in the paxillin/alpha-parvin system that may facilitate the assembly of dynamic signaling complexes in the cell.

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Cocrystal Structures of α-Parvin-CHC with Paxillin LD1, LD2, and LD4(A) Detail of the α-parvin-CHC complexes with LD1 (top), LD2 (middle), and LD4 (bottom). The left panel shows electrostatic surface renditions of α-parvin-CHC with the bound LD peptides represented by a combination of ribbon, ball-and-stick (side-chains), and cylinder (main chain) modes to indicate directionality. The right panel shows ribbon representations of α-parvin-CHC (gold) and LD peptides including those side-chains within a contact radius of 4 Å as ball-and-stick models.(B) Sequence alignment of LD peptides. Those residues ordered in the crystals are underlined; acidic residues are colored red, and basic ones are blue. Residues in contact with the protein within a radius of 4 Å are boxed. The pseudo-palindromic axis of LD1 is shown as a dashed line.
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fig3: Cocrystal Structures of α-Parvin-CHC with Paxillin LD1, LD2, and LD4(A) Detail of the α-parvin-CHC complexes with LD1 (top), LD2 (middle), and LD4 (bottom). The left panel shows electrostatic surface renditions of α-parvin-CHC with the bound LD peptides represented by a combination of ribbon, ball-and-stick (side-chains), and cylinder (main chain) modes to indicate directionality. The right panel shows ribbon representations of α-parvin-CHC (gold) and LD peptides including those side-chains within a contact radius of 4 Å as ball-and-stick models.(B) Sequence alignment of LD peptides. Those residues ordered in the crystals are underlined; acidic residues are colored red, and basic ones are blue. Residues in contact with the protein within a radius of 4 Å are boxed. The pseudo-palindromic axis of LD1 is shown as a dashed line.

Mentions: To elucidate the molecular basis for LD recognition by α-parvin-CHC, we cocrystallized α-parvin-CHC with peptides representing the high-affinity ligands LD1, LD2, and LD4. The corresponding structures at 2.1 Å, 1.8 Å, and 2.2 Å resolution, respectively, were solved by molecular replacement with the α-parvin-CHCapo structure (Table 1). In all three cases, continuous positive difference density was identified into which the peptide ligands could be placed unambiguously (Figure S3). The refined structures include residues 247–372 of α-parvin and paxillin residues 1–14, 141–155, and 262–274 for LD1, LD2, and LD4, respectively (Figures 3A and 4). The remaining C-terminal residues of the LD peptides appear disordered, presumably because they do not form contacts with α-parvin-CHC (Figure 3B). All three LD motifs bind to the same binding site on α-parvin-CHC formed by the N-linker helix, the N-terminal part of helix A and the C-terminal part of helix G, which is consistent with our results from chemical shift mapping (Figure 2). We thus conclude that the PBS region previously identified on α-parvin is not directly involved in the interaction with LD peptides.


Structural analysis of the interactions between paxillin LD motifs and alpha-parvin.

Lorenz S, Vakonakis I, Lowe ED, Campbell ID, Noble ME, Hoellerer MK - Structure (2008)

Cocrystal Structures of α-Parvin-CHC with Paxillin LD1, LD2, and LD4(A) Detail of the α-parvin-CHC complexes with LD1 (top), LD2 (middle), and LD4 (bottom). The left panel shows electrostatic surface renditions of α-parvin-CHC with the bound LD peptides represented by a combination of ribbon, ball-and-stick (side-chains), and cylinder (main chain) modes to indicate directionality. The right panel shows ribbon representations of α-parvin-CHC (gold) and LD peptides including those side-chains within a contact radius of 4 Å as ball-and-stick models.(B) Sequence alignment of LD peptides. Those residues ordered in the crystals are underlined; acidic residues are colored red, and basic ones are blue. Residues in contact with the protein within a radius of 4 Å are boxed. The pseudo-palindromic axis of LD1 is shown as a dashed line.
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fig3: Cocrystal Structures of α-Parvin-CHC with Paxillin LD1, LD2, and LD4(A) Detail of the α-parvin-CHC complexes with LD1 (top), LD2 (middle), and LD4 (bottom). The left panel shows electrostatic surface renditions of α-parvin-CHC with the bound LD peptides represented by a combination of ribbon, ball-and-stick (side-chains), and cylinder (main chain) modes to indicate directionality. The right panel shows ribbon representations of α-parvin-CHC (gold) and LD peptides including those side-chains within a contact radius of 4 Å as ball-and-stick models.(B) Sequence alignment of LD peptides. Those residues ordered in the crystals are underlined; acidic residues are colored red, and basic ones are blue. Residues in contact with the protein within a radius of 4 Å are boxed. The pseudo-palindromic axis of LD1 is shown as a dashed line.
Mentions: To elucidate the molecular basis for LD recognition by α-parvin-CHC, we cocrystallized α-parvin-CHC with peptides representing the high-affinity ligands LD1, LD2, and LD4. The corresponding structures at 2.1 Å, 1.8 Å, and 2.2 Å resolution, respectively, were solved by molecular replacement with the α-parvin-CHCapo structure (Table 1). In all three cases, continuous positive difference density was identified into which the peptide ligands could be placed unambiguously (Figure S3). The refined structures include residues 247–372 of α-parvin and paxillin residues 1–14, 141–155, and 262–274 for LD1, LD2, and LD4, respectively (Figures 3A and 4). The remaining C-terminal residues of the LD peptides appear disordered, presumably because they do not form contacts with α-parvin-CHC (Figure 3B). All three LD motifs bind to the same binding site on α-parvin-CHC formed by the N-linker helix, the N-terminal part of helix A and the C-terminal part of helix G, which is consistent with our results from chemical shift mapping (Figure 2). We thus conclude that the PBS region previously identified on α-parvin is not directly involved in the interaction with LD peptides.

Bottom Line: Cocrystal structures with these LD motifs reveal the molecular details of their interactions with a common binding site on alpha-parvin-CH(C), which is located at the rim of the canonical fold and includes part of the inter-CH domain linker.Surprisingly, this binding site can accommodate LD motifs in two antiparallel orientations.Taken together, these results reveal an unusual degree of binding degeneracy in the paxillin/alpha-parvin system that may facilitate the assembly of dynamic signaling complexes in the cell.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Molecular Biophysics, University of Oxford, Oxford OX1 3QU, United Kingdom.

ABSTRACT
The adaptor protein paxillin contains five conserved leucine-rich (LD) motifs that interact with a variety of focal adhesion proteins, such as alpha-parvin. Here, we report the first crystal structure of the C-terminal calponin homology domain (CH(C)) of alpha-parvin at 1.05 A resolution and show that it is able to bind all the LD motifs, with some selectivity for LD1, LD2, and LD4. Cocrystal structures with these LD motifs reveal the molecular details of their interactions with a common binding site on alpha-parvin-CH(C), which is located at the rim of the canonical fold and includes part of the inter-CH domain linker. Surprisingly, this binding site can accommodate LD motifs in two antiparallel orientations. Taken together, these results reveal an unusual degree of binding degeneracy in the paxillin/alpha-parvin system that may facilitate the assembly of dynamic signaling complexes in the cell.

Show MeSH
Related in: MedlinePlus