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 be tween 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 dystrop hy. Previously, the cysteine-rich region and first half of the carboxy-term inal domain of dystrophin were shown to interact with beta-dystroglycan thr ough a stretch of fifteen amino acids at the carboxy-terminus of beta-dystr oglycan. This region of dystrophin implicated in binding beta-dystroglycan contains four modular protein domains: a WW domain, two putative Ca2+-bindi ng 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 t o bind ligands that contain a Pro-Pro-x-Tyr core motif (where x is any amin o acid). Here we elucidate the role of the WW domain of dystrophin and surr ounding sequence in binding beta-dystroglycan. We show that the WW domain o f 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 o f beta-dystroglycan occurs via a beta-dystroglycan Pro-Pro-x-Tyr core motif . Targeted mutagenesis of conserved WW domain residues reveals that the dys trophin/beta-dystroglycan interaction occurs primarily through the WW domai n of dystrophin. Precise mapping of this interaction could aid in therapeut ic design.