ZZ domain of dystrophin and utrophin: topology and mapping of a beta-dystroglycan interaction site.
Dystrophin forms part of a vital link between actin cytoskeleton and extracellular matrix via the transmembrane adhesion receptor dystroglycan. Dystrophin and its autosomal homologue utrophin interact with beta-dystroglycan via their highly conserved C-terminal cysteine-rich regions, comprising the WW domain (protein-protein interaction domain containing two conserved tryptophan residues), EF hand and ZZ domains. The EF hand region stabilizes the WW domain providing the main interaction site between dystrophin or utrophin and dystroglycan. The ZZ domain, containing a predicted zinc finger motif, stabilizes the WW and EF hand domains and strengthens the overall interaction between dystrophin or utrophin and beta-dystroglycan. Using bacterially expressed ZZ domain, we demonstrate a conformational effect of zinc binding to the ZZ domain, and identify two zinc-binding regions within the ZZ domain by SPOTs overlay assays. Epitope mapping of the dystrophin ZZ domain was carried out with new monoclonal antibodies by ELISA, overlay assay and immunohistochemistry. One monoclonal antibody defined a discrete region of the ZZ domain that interacts with beta-dystroglycan. The epitope was localized to the conformationally sensitive second zinc-binding site in the ZZ domain. Our results suggest that residues 3326-3332 of dystrophin form a crucial part of the contact region between dystrophin and beta-dystroglycan and provide new insight into ZZ domain organization and function.
Hnia
Karim
K
Université Montpellier 1, Unité de Formation et de Recherche de Médecine, Laboratoire de Physiologie des Interactions, Institut de Biologie, Boulevard Henri IV, F-34062, France.
Zouiten
Dora
D
Cantel
Sonia
S
Chazalette
Delphine
D
Hugon
Gérald
G
Fehrentz
Jean-Alain
JA
Masmoudi
Ahmed
A
Diment
Ann
A
Bramham
Janice
J
Mornet
Dominique
D
Winder
Steve J
SJ
D000595:Amino Acid Sequence; D001665:Binding Sites; D049030:Dystroglycans / Q000737:chemistry* / Q000378:metabolism; D016189:Dystrophin / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D020125:Mutation, Missense; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D049411:Utrophin / Q000737:chemistry* / Q000378:metabolism; D015032:Zinc / Q000737:chemistry / Q000378:metabolism
D016189:Dystrophin; D049411:Utrophin; D049030:Dystroglycans; D015032:Zinc