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Structural Features of the Full-Length Adaptor Protein GADS in Solution Determined Using Small-Angle X-Ray Scattering

Oscar Moran ^*, Manfred W. Roessle , Roy A. Mariuzza  and Nazzareno Dimasi 

^* Istituto di Biofisica, Consiglio Nazionale delle Ricerche, 16149 Genova, Italy; European Molecular Biology Laboratory, Hamburg Outstation, D-22603 Hamburg, Germany; Center for Advanced Research in Biotechnology, 9600 Gudelsky Drive, 20870 Rockville, Maryland; and Istituto Giannina Gaslini, 16147 Genova, Italy

Correspondence: Address reprint requests to Oscar Moran, Istituto di Biofisica, Consiglio Nazionale delle Ricerche, 16149 Genova, Italy. E-mail: moran{at}ge.ibf.cnr.it; and to Nazzareno Dimasi, Istituto Giannina Gaslini, 16147 Genova, Italy. E-mail: ndimasi{at}gmail.com.

The Grb2-related adaptor protein GADS plays a central role during the initial phases of signal transduction in T lymphocytes. GADS possesses N- and C-terminal Src homology 3 (SH3) domains flanking a central Src homology 2 (SH2) domain and a 126-residue region rich in glutamine and proline residues, presumed to be largely unstructured. The SH2 domain of GADS binds the adaptor protein LAT; the C-terminal SH3 domain pairs GADS to the adaptor protein SLP-76, whereas the function of the central region is unknown. High-resolution three-dimensional models are available for the isolated SH2 and C-terminal SH3 domains in complex with their respective binding partners, LAT and SLP-76. However, in part because of its intrinsic instability, there is no structural information for the entire GADS molecule. Here, we report the low-resolution structure of full-length GADS in solution using small-angle x-ray scattering (SAXS). Based on the SAXS data, complemented by gel filtration experiments, we show that full-length GADS is monomeric in solution and that its overall structural parameters are smaller than those expected for a protein with a long unstructured region. Ab initio and rigid body modeling of the SAXS data reveal that full-length GADS is a relatively compact molecule and that the potentially unstructured region retains a significant degree of structural order. The biological function of GADS is discussed based on its overall structure.