Contribution of the different modules in the utrophin carboxy-terminal region to the formation and regulation of the DAP complex

The carboxy-terminal region of utrophin, like the homologous proteins dystr ophin, Drp2 and dystrobrevins, contains structural domains frequently invol ved in protein-protein interaction. These domains (WW, EF hands, ZZ and H1- H2) mediate recognition and binding to a multicomponent complex of proteins , also known as dystrophin-associated proteins (DAPs) for their association with dystrophin, the product of the gene, mutated in Duchenne muscular dys trophy. We have exploited phage display and in vitro binding assays to stud y the recognition specificity of the different domains of the utrophin carb oxy-terminus. We found that none of the carboxy-terminal domains of utrophi n, when isolated from its structural contest, selects specific ligand pepti des from a phage-displayed peptide library. By contrast, panning with an ex tended region containing the WW, EF hands, and ZZ domain defines the consen sus binding motif, PPxY which is also found in beta-dystroglycan, a compone nt of the DAP complex that interacts with utrophin in several tissues, WW-m ediated binding to PPxY peptides and to beta-dystroglycan requires the pres ence of the EF hands and ZZ domain. When the ZZ domain is either deleted or engaged in binding to calmodulin, the utrophin beta-dystroglycan complex c annot be formed. These findings suggest a potential regulatory mechanism bg l means of which the attachment of utrophin to the DAP complex can he modul ated by the Ca2+-dependent binding of calmodulin. The remaining two motifs found in the carboxy-terminus (H1-H2) mediate the formation of utrophin-dys trobrevin hybrids hut do not select ligands in a repertoire of random nonap eptides. (C) 2000 Federation of European Biochemical Societies.