The molecular basis of alpha-dystroglycan hypoglycosylation: a crystallographic and SAXS study

Dystroglycan (DG) is a glycoprotein complex that links the cytoskeleton with the extracellular matrix. DG is composed of two subunits: alpha-DG and beta-DG. alpha-DG is a highly glycosylated extracellular protein which is essential for high-affinity binding of extracellular matrix proteins such as laminins. The hypoglycosylation of alpha-DG weakens this interaction affinity resulting in severe pathological states. The N-terminal region (aa 50-313 in mouse) of alpha-DG plays a crucial role in the glycosylation of the ?-DG mucin-like domain, being required by the glycosyltransferase LARGE during the extension of the O-glycans implicated in laminin binding. Furthermore, pathological missense mutations, observed at the N-terminal region of alpha-DG, are responsible of alpha-DG hypoglycosylation states resulting in primary distroglycanopathies1. We have been investigating the structures of the three point mutants V72I (V72I-ma-DG), D109N (D109N-ma-DG) and T190M2 (T190M-ma-DG) of mouse alpha-DG (ma-DG), combining X-ray crystallography and in solution Small Angle X-ray Scattering (SAXS). The purpose of this study is to gain evidences about the structural determinants of N-terminal ?-DG that are functionally relevant for its glycosylation pathway. The crystal structures of the three mutants display the same overall structure of WT-ma-DG. In contrast, the solution structural models obtained by SAXS analysis depict a different scenario, where the WT-a-DG is quite flexible in solution, assuming more than one conformation. The comparison of SAXS data from WT-ma-DG and its mutants demonstrates the presence of a perturbation, in both the conformations as well as in the partition among different populations, which is mediated by the flexible peptide linking the Ig-like and S6 small-ribosomal domains, embodied in the alpha-DG N-terminal region. SAXS data thus suggest a more complex and dynamical situation with respect to the crystallographic evidences that may have functional implications for DG post-translational processing and for the interactions with its binding partners.