RESUMEN
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.
Asunto(s)
Actinina/química , Actinina/metabolismo , Paxillin/química , Paxillin/metabolismo , Secuencias de Aminoácidos , Secuencia de Aminoácidos , Humanos , Proteínas Repetidas Ricas en Leucina , Proteínas de Microfilamentos , Modelos Moleculares , Datos de Secuencia Molecular , Unión Proteica , Mapeo de Interacción de Proteínas , Proteínas/química , Proteínas/metabolismo , Homología de Secuencia de Aminoácido , Transducción de Señal/fisiologíaRESUMEN
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.
Asunto(s)
Proteínas Adaptadoras Transductoras de Señales , Estructura Secundaria de Proteína , Estructura Terciaria de Proteína , Proteínas Tirosina Quinasas/química , Proteínas Tirosina Quinasas/metabolismo , Transducción de Señal/fisiología , Secuencia de Aminoácidos , Animales , Sitios de Unión , Adhesión Celular/fisiología , Moléculas de Adhesión Celular/química , Moléculas de Adhesión Celular/metabolismo , Cristalografía por Rayos X , Proteínas del Citoesqueleto/química , Proteínas del Citoesqueleto/metabolismo , Quinasa 1 de Adhesión Focal , Proteína-Tirosina Quinasas de Adhesión Focal , Proteína Adaptadora GRB2 , Humanos , Modelos Moleculares , Datos de Secuencia Molecular , Paxillin , Fosfoproteínas/química , Fosfoproteínas/metabolismo , Proteínas Tirosina Quinasas/genética , Proteínas/metabolismo , Alineación de Secuencia , Tirosina/metabolismo , Dominios Homologos srcRESUMEN
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.