Sequence analysis suggests the presence of an IG-like domain in the N-terminal region of alpha-dystroglycan which was crystallized after mutation of a protease susceptible site (Arg168-->His).
Recently, we demonstrated that the N-terminal region of mouse alpha-dystroglycan represents an autonomously folding globular domain, organized into at least two subdomains (Brancaccio et al., Eur. J. Biochem. 246, 166-172, 1997). We have now found a similarity between a part of the alpha-dystroglycan N-terminal sequence (approximately from position 80 to 180) and several protein sequences belonging to the immunoglobulin kappa family. Moreover, we have recombinantly expressed and purified a 31 kDa protein fragment which matches the entire alpha-dystroglycan N-terminal globular domain. To prevent the action of bacterial endogenous proteases and/or thrombin, which cleaves the protein into two fragments at an Arg-Ala trypsin-sensitive site in positions 168-169, we have introduced a single mutation (Arg168-->His), thus making the whole domain more stable and suitable for crystallization. Crystals of this mutant protein were obtained by vapor diffusion using the hanging drop technique, and they diffract to 0.28 nm Bragg spacing.
Bozic
D
D
University Biocenter, Biophysical Chemistry, University of Basel, Switzerland.
Engel
J
J
Brancaccio
A
A
D000595:Amino Acid Sequence; D000818:Animals; D001120:Arginine; D001665:Binding Sites; D003460:Crystallization; D003598:Cytoskeletal Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D049030:Dystroglycans; D010450:Endopeptidases / Q000378:metabolism*; D006639:Histidine; D007145:Immunoglobulin kappa-Chains / Q000737:chemistry; D008562:Membrane Glycoproteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D008969:Molecular Sequence Data; D009154:Mutation; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D014961:X-Ray Diffraction
D003598:Cytoskeletal Proteins; D007145:Immunoglobulin kappa-Chains; D008562:Membrane Glycoproteins; D011994:Recombinant Proteins; D049030:Dystroglycans; D006639:Histidine; D001120:Arginine; D010450:Endopeptidases