Spatial arrangement of LD motif-interacting residues on focal adhesion targeting domain of Focal Adhesion Kinase determine domain-motif interaction affinity and specificity
Resumen: Background: Leucine rich Aspartate motifs (LD motifs) are molecular recognition motifs on Paxillin that recognize LD-motif binding domains (LDBD) of a number of focal adhesion proteins in order to carry out downstream signaling and actin cytoskeleton remodeling. In this study, we identified structural features within LDBDs that influence their binding affinity with Paxillin LD motifs. Methods: Various point mutants of focal adhesion targeting (FAT) domain of Focal Adhesion Kinase (FAK) were created by moving a key Lysine residue two and three helical turns in order to match the unique conformations as observed in LDBDs of two other focal adhesion proteins, Vinculin and CCM3. Results: This led to identify a mutant of FAT domain of FAK, named as FAT(NV) (Asn992 of FAT domain was replaced by Val), with remarkable high affinity for LD1 (Kd = 1.5 µM vs no-binding with wild type) and LD2 peptides (Kd = 7.2 µM vs 63 µM with wild type). Consistently, the focal adhesions of MCF7 cells expressing FAK(NV) were highly stable (turnover rate = 1.25 × 10-5 µm2/s) as compared to wild type FAK transfected cells (turnover rate = 1.5 × 10-3 µm2/s). Conclusions: We observed that the relative disposition of key LD binding amino-acids at LDBD surface, hydrophobic burial of long Leucine side chains of LD-motifs and complementarity of charged surfaces are the key factors determining the binding affinities of LD motifs with LDBDs. General significance: Our study will help in protein engineering of FAT domain of FAK by modulating FAK-LD motif interactions which have implications in cellular focal adhesions and cell migration.
Idioma: Inglés
DOI: 10.1016/j.bbagen.2019.129450
Año: 2020
Publicado en: BIOCHIMICA ET BIOPHYSICA ACTA-GENERAL SUBJECTS 1864, 1 (2020), 129450 1-9
ISSN: 0304-4165

Originalmente disponible en: Texto completo de la revista

Factor impacto JCR: 3.77 (2020)
Categ. JCR: BIOPHYSICS rank: 25 / 71 = 0.352 (2020) - Q2 - T2
Categ. JCR: BIOCHEMISTRY & MOLECULAR BIOLOGY rank: 154 / 297 = 0.519 (2020) - Q3 - T2

Factor impacto SCIMAGO: 1.204 - Biochemistry (Q1) - Molecular Biology (Q1) - Biophysics (Q1)

Tipo y forma: Article (Published version)
Área (Departamento): Área Bioquímica y Biolog.Mole. (Dpto. Bioq.Biolog.Mol. Celular)

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