title: Structural Basis for Noncanonical Substrate Recognition of Cofilin/ADF Proteins by LIM Kinases.
abstract: Cofilin/actin-depolymerizing factor (ADF) proteins are critical nodes that relay signals from protein kinase cascades to the actin cytoskeleton, in particular through site-specific phosphorylation at residue Ser3. This is important for regulation of the roles of cofilin in severing and stabilizing actin filaments. Consequently, cofilin/ADF Ser3 phosphorylation is tightly controlled as an almost exclusive substrate for LIM kinases. Here we determine the LIMK1:cofilin-1 co-crystal structure. We find an interface that is distinct from canonical kinase-substrate interactions. We validate this previously unobserved mechanism for high-fidelity kinase-substrate recognition by in vitro kinase assays, examination of cofilin phosphorylation in mammalian cells, and functional analysis in S. cerevisiae. The interface is conserved across all LIM kinases. Remarkably, we also observe both pre- and postphosphotransfer states in the same crystal lattice. This study therefore provides a molecular understanding of how kinase-substrate recognition acts as a gatekeeper to regulate actin cytoskeletal dynamics.
authors.affiliation: Department of Pharmacology, Yale University School of Medicine, New Haven, CT 06520, USA.
authors.forename: Stephanie
authors.initials: S
authors.lastname: Hamill
authors.affiliation: Department of Pharmacology, Yale University School of Medicine, New Haven, CT 06520, USA.
authors.forename: Hua Jane
authors.initials: HJ
authors.lastname: Lou
authors.affiliation: Department of Pharmacology, Yale University School of Medicine, New Haven, CT 06520, USA; Yale Cancer Center, Yale University School of Medicine, New Haven, CT 06520, USA.
authors.forename: Benjamin E
authors.initials: BE
authors.lastname: Turk
authors.affiliation: Department of Pharmacology, Yale University School of Medicine, New Haven, CT 06520, USA; Department of Molecular Biophysics and Biochemistry, Yale University School of Medicine, New Haven, CT 06520, USA; Yale Cancer Center, Yale University School of Medicine, New Haven, CT 06520, USA. Electronic address: titus.boggon@yale.edu.
authors.forename: Titus J
authors.initials: TJ
authors.lastname: Boggon
mesh_terms: D001665:Binding Sites; D020134:Catalytic Domain; D051338:Cofilin 1 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D018360:Crystallography, X-Ray; D057809:HEK293 Cells; D006801:Humans; D054461:Lim Kinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D009154:Mutation; D010766:Phosphorylation; D011485:Protein Binding; D011487:Protein Conformation; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012694:Serine; D013329:Structure-Activity Relationship; D013379:Substrate Specificity; D014162:Transfection
chemical_list: C495810:CFL1 protein, human; C495813:COF1 protein, S cerevisiae; D051338:Cofilin 1; D029701:Saccharomyces cerevisiae Proteins; D012694:Serine; C515892:LIMK1 protein, human; D054461:Lim Kinases