The effect of an ionic detergent on the natively unfolded β‐dystroglycan ectodomain and on its interaction with α‐dystroglycan

Dystroglycan (DG) is an adhesion complex, expressed in a wide variety of tissues, formed by an extracellular and a transmembrane subunit, α‐DG and β‐DG, respectively, interacting noncovalently. Recently, we have shown that the recombinant ectodomain of β‐DG, β‐DG(654–750), behaves as a natively unfolded protein, as it is able to bind the C‐terminal domain of α‐DG, while not displaying a defined structural organization. We monitored the effect of a commonly used denaturing agent, the anionic detergent sodium dodecylsulphate (SDS), on β‐DG(654–750) using a number of biophysical techniques. Very low concentrations of SDS (≤2 mM) affect both tryptophan fluorescence and circular dichroism of β‐DG, and significantly perturb the interaction with the α‐DG subunit as shown by solid‐phase binding assays and fluorescence titrations in solution. This result confirms, as recently proposed for natively unfolded proteins, that β‐DG(654–750) exists in a native state, which is crucial to fulfill its biological function. Two‐dimensional NMR analysis shows that SDS does not induce any evident conformational rearrangement within the ectodomain of β‐DG. Its first 70 amino acids, which show a lower degree of mobility, interact with the detergent, but this does not change the amount of secondary structure, whereas the highly flexible and mobile C‐terminal region of β‐DG(654–750) remains largely unaffected, even at a very high SDS concentration (up to 50 mM). Our data indicate that SDS can be used as a useful tool for investigating natively unfolded proteins, and confirm that the β‐DG ectodomain is an interesting model system.