RESUMO
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.
Assuntos
Actinina/química , Actinina/metabolismo , Paxilina/química , Paxilina/metabolismo , Motivos de Aminoácidos , Sequência de Aminoácidos , Humanos , Proteínas de Repetições Ricas em Leucina , Proteínas dos Microfilamentos , Modelos Moleculares , Dados de Sequência Molecular , Ligação Proteica , Mapeamento de Interação de Proteínas , Proteínas/química , Proteínas/metabolismo , Homologia de Sequência de Aminoácidos , Transdução de Sinais/fisiologiaRESUMO
The localization of focal adhesion kinase (FAK) to sites of integrin clustering initiates downstream signaling. The C-terminal focal adhesion targeting (FAT) domain causes this localization by interacting with talin and paxillin. FAT also mediates signaling through Grb2 via phosphorylated Y925. We report two crystal structures of the FAT domain. Large rearrangements of the structure are indicated to allow phosphorylation of Y925 and subsequent interaction with Grb2. Sequence homology and structural compatibility suggest a FAT-like fold for the C-terminal domains of CAS, Efs/Sin, and HEF1. A structure-based alignment including these proteins and the vinculin tail domain reveals a conserved region that could play a role in focal adhesion targeting. Previously postulated "paxillin binding subdomains" may contribute to structural integrity rather than directly to paxillin binding.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal , Estrutura Secundária de Proteína , Estrutura Terciária de Proteína , Proteínas Tirosina Quinases/química , Proteínas Tirosina Quinases/metabolismo , Transdução de Sinais/fisiologia , Sequência de Aminoácidos , Animais , Sítios de Ligação , Adesão Celular/fisiologia , Moléculas de Adesão Celular/química , Moléculas de Adesão Celular/metabolismo , Cristalografia por Raios X , Proteínas do Citoesqueleto/química , Proteínas do Citoesqueleto/metabolismo , Quinase 1 de Adesão Focal , Proteína-Tirosina Quinases de Adesão Focal , Proteína Adaptadora GRB2 , Humanos , Modelos Moleculares , Dados de Sequência Molecular , Paxilina , Fosfoproteínas/química , Fosfoproteínas/metabolismo , Proteínas Tirosina Quinases/genética , Proteínas/metabolismo , Alinhamento de Sequência , Tirosina/metabolismo , Domínios de Homologia de srcRESUMO
Focal adhesions (FAs) are large submembrane signaling complexes formed at sites of cellular attachment to the extracellular matrix. The interaction of LD motifs with their targets plays an important role in the assembly of FAs. We have determined the molecular basis for the recognition of two paxillin LD motifs by the FA targeting (FAT) domain of FA kinase using a combination of X-ray crystallography, solution NMR, and homology modeling. The four-helix FAT domain displays two LD binding sites on opposite sites of the molecule that bind LD peptides in a helical conformation. Threading studies suggest that the LD-interacting domain of p95PKL shares a common four-helical core with the FAT domain and the tail of vinculin, defining a structural family of LD motif binding modules.