Enzymatic processing by MMP-2 and MMP-9 of wild-type and mutated mouse ss-dystroglycan

Dystroglycan (DG) is a membrane-associated protein complex formed by two noncovalently linked subunits, a-DG, a highly glycosylated extracellular protein, and beta-DG, a transmembrane protein. The interface between the two DG subunits, which is crucial to maintain the integrity of the plasma membrane, involves the C-terminal domain of a-DG and the N-terminal extracellular domain of beta-DG. It is well known that under both, physiological and pathological conditions, gelatinases (i.e. MMP-9 and/or MMP-2) can degrade DG, disrupting the connection between the extracellular matrix and the cytoskeleton. However, the molecular mechanisms and the exact cleavage sites underlying these events are still largely unknown. In a previous study, we have characterized the enzymatic digestion of the murine beta-DG ectodomain by gelatinases, identifying a main cleavage site on the beta-DG ectodomain produced by MMP-9. In this article, we have deepened the pattern of the beta-DG ectodomain digestion by MMP-2 by using a combined approach based on SDS-PAGE, Orbitrap, and HPLC-ESI-IT mass spectrometry. Furthermore, we have characterized the kineticparameters of the digestion of some beta-DG ectodomain mutants by gelatinases.