The WW domain of dystrophin requires EF-hands region to interact with beta-dystroglycan.
Skeletal muscle dystrophin is a 427 kDa protein thought to act as a link between the actin cytoskeleton and the extracellular matrix. Perturbations of the dystrophin-associated complex, for example, between dystrophin and the transmembrane glycoprotein beta-dystroglycan, may lead to muscular dystrophy. Previously, the cysteine-rich region and first half of the carboxy-terminal domain of dystrophin were shown to interact with beta-dystroglycan through a stretch of fifteen amino acids at the carboxy-terminus of beta-dystroglycan. This region of dystrophin implicated in binding beta-dystroglycan contains four modular protein domains: a WW domain, two putative Ca2+-binding EF-hand motifs, and a putative zinc finger ZZ domain. The WW domain is a globular domain of 38-40 amino acids with two highly conserved tryptophan residues spaced 20-22 amino acids apart. A subset of WW domains was shown to bind ligands that contain a Pro-Pro-x-Tyr core motif (where x is any amino acid). Here we elucidate the role of the WW domain of dystrophin and surrounding sequence in binding beta-dystroglycan. We show that the WW domain of dystrophin along with the EF-hand motifs binds to the carboxy-terminus of beta-dystroglycan. Through site-specific mutagenesis and in vitro binding assays, we demonstrate that binding of dystrophin to the carboxy-terminus of beta-dystroglycan occurs via a beta-dystroglycan Pro-Pro-x-Tyr core motif. Targeted mutagenesis of conserved WW domain residues reveals that the dystrophin/beta-dystroglycan interaction occurs primarily through the WW domain of dystrophin. Precise mapping of this interaction could aid in therapeutic design.
Rentschler
S
S
Department of Biochemistry, Mount Sinai School of Medicine, New York, NY 10029, USA.
Linn
H
H
Deininger
K
K
Bedford
M T
MT
Espanel
X
X
Sudol
M
M
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002478:Cells, Cultured; D003598:Cytoskeletal Proteins / Q000235:genetics / Q000378:metabolism*; D017931:DNA Primers; D049030:Dystroglycans; D016189:Dystrophin / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D008562:Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D008969:Molecular Sequence Data; D011233:Precipitin Tests; D011485:Protein Binding; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism
D003598:Cytoskeletal Proteins; C483081:DAG1 protein, human; D017931:DNA Primers; D016189:Dystrophin; D008562:Membrane Glycoproteins; D011993:Recombinant Fusion Proteins; D049030:Dystroglycans