title: Identification of N- and C-terminal residues involved in MAP kinase phosphatase 3 (MKP3) interdomain binding and auto-inhibition.
abstract: Interdomain binding has been shown to play an important role in the regulation of MAP kinase phosphatase 3 (MKP3), a phosphatase involved in control of ERK signalling pathways. In this study the residues in N- and C-terminal domains responsible for MKP3 interdomain binding are identified. Peptides from the N-terminal substrate-binding domain of MKP3 were assessed for their ability to bind the C-terminal catalytic domain using surface plasmon resonance. The data indicate that the residues 77-97 (the Post-KIM peptide) in the MKP3 N-terminal domain are responsible for its binding to the C-terminal catalytic domain. Residues in the C-terminal domain that might be important to interdomain binding were identified using data in the existing literature. Variants in which these residues had been altered were examined by circular dichroism and enzymatic assays to ensure retention of their structure and catalytic properties before being assessed for their ability to bind the Post-KIM peptide. The data show that glutamic acid 248, asparagine 267 and, to a lesser extent, arginine 299 are important for the interaction between the MKP3 C-terminal and the N-terminal domains. The identified residues map to a region on the surface of the C-terminal domain that appears complementary to the N-terminal domain surface defined by the Post-KIM peptide. This interdomain binding site is distinct from the substrate interaction sites.
authors.affiliation: Centre for Biologics Research, Biologics and Genetic Therapies Directorate, Health Canada, 251 Sir Frederick Banting Driveway, Ottawa, Ontario K1A 0K9, Canada.
authors.forename: John K
authors.initials: JK
authors.lastname: Mark
authors.forename: Michael J W
authors.initials: MJ
authors.lastname: Johnston
authors.forename: Mary Alice
authors.initials: MA
authors.lastname: Hefford
mesh_terms: D020134:Catalytic Domain; D002942:Circular Dichroism; D054642:Dual Specificity Phosphatase 6 / Q000737:chemistry* / Q000378:metabolism*; D008958:Models, Molecular; D011485:Protein Binding; D017510:Protein Folding
chemical_list: D054642:Dual Specificity Phosphatase 6