title: DEF pocket in p38α facilitates substrate selectivity and mediates autophosphorylation.
abstract: Signaling processes are primarily promoted by molecular recognition and corresponding protein-protein interactions. One of the key eukaryotic signaling pathways is the MAP kinase cascade involved in vital cellular processes such as cell proliferation, differentiation, apoptosis, and stress response. The principle recognition site of MAP kinases, the common docking (CD) region, forms selective interactions with substrates, upstream activators, and phosphatases. A second docking site, defined as the DEF site interaction pocket (DEF pocket), is formed subsequent to ERK2 and p38α activation. Both crystal structures of p38α in its dually phosphorylated form and of intrinsically active mutants showed the DEF pocket, giving motivation for studying its role in substrate activation and selectivity. Mutating selected DEF pocket residues significantly decreased the phosphorylation levels of three p38α substrates (ATFII, Elk-1, and MBP) with no apparent effect on the phosphorylation of MK2 kinase. Conversely, mutating the CD region gave the opposite effect, suggesting p38α substrates can be classified into DEF-dependent and DEF-independent substrates. In addition, mutating DEF pocket residues decreased the autophosphorylation capability of intrinsically active p38α mutants, suggesting DEF-mediated trans-autophosphorylation in p38α. These results could contribute to understanding substrate selectivity of p38α and serve as a platform for designing p38α-selective DEF site blockers, which partially inhibit p38α binding DEF-dependent substrates, whereas maintaining its other functions intact. In this context, preliminary results using synthetic peptides reveal significant inhibition of substrate phosphorylation by activated p38α.
authors.affiliation: Department of Biological Chemistry, The Alexander Silverman Institute of Life Sciences, The Hebrew University of Jerusalem, Jerusalem 91904, Israel.
authors.forename: Netanel
authors.initials: N
authors.lastname: Tzarum
authors.forename: Nadav
authors.initials: N
authors.lastname: Komornik
authors.forename: Dorin
authors.initials: D
authors.lastname: Ben Chetrit
authors.forename: David
authors.initials: D
authors.lastname: Engelberg
authors.forename: Oded
authors.initials: O
authors.lastname: Livnah
mesh_terms: D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D006801:Humans; D019950:Mitogen-Activated Protein Kinase 1 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D048308:Mitogen-Activated Protein Kinase 14 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D009154:Mutation; D010455:Peptides / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation / Q000502:physiology; D013379:Substrate Specificity
chemical_list: D010455:Peptides; C535150:MAPK1 protein, human; D019950:Mitogen-Activated Protein Kinase 1; D048308:Mitogen-Activated Protein Kinase 14
keywords: Autophosphorylation; Docking Region; MAP Kinase; MAP Kinases (MAPKs); Molecular Docking; Protein Phosphorylation; Substrate Selectivity; Threonine-serine Protein Kinase; p38