Structure of the N-WASP EVH1 Domain-WIP
Complex: Insight into the Molecular Basis
of Wiskott-Aldrich Syndrome
Supplemental Figure S1. Presence of Covalent Linker Does Not Effect WIP-EVH1
Complex Structure
(A) In addition to the covalently linked construct used for the bulk of these
studies, we also generated a construct containing a thrombin cleavage site
between the WIP peptide and the EVH1 domain. Samples were prepared and the
second construct cleaved to completion as described in Experimental Procedures.
(B) 2D 15N-1H HSQC spectra of the covalently tethered complex (black) and
the cleaved complex (red). Bottom image shows an overlay of the two spectra.
In all images EVH1 domain residues at the binding surface are labeled in violet
and WIP peptide residues are labeled in gold. Labeled EVH1 residues include
those that that bind the N-terminal part of the peptide (Y46, W56, F104, T106;
see Figure 5) and those that bind near the C terminus of the peptide (E90,
Y92, G109, C112). Visible peptide residues range from 461 to 480. The spectra
are virtually identical except for minor shifts (green) at residues near the
cleavage termini. A few new resonances in the cleaved spectrum correspond
to the new residues introduced by the thrombin site.
Conclusion: the structure of the EVH1-WIP complex remains the same even after
cleavage of the covalent tether.
