Structural determinants of the C-terminal helix-kink-helix motif essential for protein stability and survival promoting activity of DJ-1.

Full Abstract

Mutations in the PARK7 gene encoding DJ-1 cause autosomal recessive Parkinson disease. The most deleterious point mutation is the L166P substitution, which resides in a structure motif comprising two alpha-helices (G and H) separated by a kink. Here we subjected the C-terminal helix-kink-helix motif to systematic site-directed mutagenesis, introducing helix-incompatible proline residues as well as conservative substitutions into the helical interface. Furthermore, we generated deletion mutants lacking the H-helix, the kink, and the entire C terminus. When transfected into neural and nonneural cell lines, steady-state levels of G-helix breaking and kink deletion mutants were dramatically lower than wild-type DJ-1. The effects of H-helix breakers were comparably smaller, and the non-helix breaking mutants only slightly destabilized DJ-1. The decreased steady-state levels were due to accelerated protein degradation involving in part the proteasome. G-helix breaking DJ-1 mutations abolished dimer formation. These structural perturbations had functional consequences on the cytoprotective activities of DJ-1. The destabilizing mutations conferred reduced cytoprotection against H(2)O(2) in transiently retransfected DJ-1 knock-out mouse embryonic fibroblasts. The loss of survival promoting activity of the DJ-1 mutants with destabilizing C-terminal mutations correlated with impaired anti-apoptotic signaling. We found that wild-type, but not mutant DJ-1 facilitated the Akt pathway and simultaneously blocked the apoptosis signal-regulating kinase 1, with which DJ-1 interacted in a redox-dependent manner. Thus, the G-helix and kink are critical determinants of the C-terminal helix-kink-helix motif, which is absolutely required for stability and the regulation of survival-promoting redox signaling of the Parkinson disease-associated protein DJ-1.

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Author information

Author/s: Görner, Karin (K); Holtorf, Eve (E); Waak, Jens (J); Pham, Thu-Trang (TT); Vogt-Weisenhorn, Daniela M (DM); Wurst, Wolfgang (W); Haass, Christian (C); Kahle, Philipp J (PJ);

Affiliation: Laboratory of Alzheimer's and Parkinson's Disease Research, Department of Biochemistry, Ludwig Maximilians University of Munich, 80336 Munich, Germany.

Journal and publication information

Publication Type: Journal Article; Research Support, Non-U.S. Gov't

Journal: The Journal of biological chemistry (J Biol Chem), published in United States. (Language: eng)

Reference: 2007-May; vol 282 (issue 18) : pp 13680-91

Dates: Created 2007/04/30; Completed 2007/06/14; Revised 2007/11/15;

PMID: 17331951, status: MEDLINE (last retrieval date: 12/26/2008)

Sourced from the National Library of Medicine. Abstract text and other information may be subject to copyright.

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MeSH headings (categories)

This article was linked to the MESH Headings shown below.

Amino Acid Motifs - genetics
Animals
Apoptosis - drug effects; genetics
Cell Survival - drug effects; genetics
Cytoprotection - drug effects; genetics
Embryo, Mammalian - metabolism
Fibroblasts - metabolism
Humans
Hydrogen Peroxide - pharmacology
Intracellular Signaling Peptides and Proteins - genetics
MAP Kinase Kinase Kinase 5 - genetics; metabolism
Mice
Mice, Knockout

Mutagenesis, Site-Directed
Mutation, Missense
Oncogene Proteins - biosynthesis; genetics
Oxidants - pharmacology
PC12 Cells
Parkinson Disease - genetics; metabolism
Protein Folding
Protein Processing, Post-Translational - drug effects; genetics
Protein Structure, Tertiary - genetics
Proto-Oncogene Proteins c-akt - genetics; metabolism
Rats
Signal Transduction - drug effects; genetics
Structure-Activity Relationship

Associated Chemicals: DJ-1 protein, mouse (0) ; Intracellular Signaling Peptides and Proteins (0) ; Oncogene Proteins (0) ; Oxidants (0) ; PARK7 protein, human (0) ; Hydrogen Peroxide (7722-84-1) ; MAP Kinase Kinase Kinase 5 (EC 2.7.1.37) ; Proto-Oncogene Proteins c-akt (EC 2.7.1.37)

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