SAXS study of the PIR domain from the Grb14 molecular adaptor: a natively unfolded protein with a transient structure primer?
Grb14 belongs to the Grb7 family of adapters and was identified as a negative regulator of insulin signal transduction. Between the PH (pleckstrin homology) and SH2 (Src homology 2) domains is a new binding domain implicated in the interaction with receptor tyrosine kinases called PIR (phosphorylated insulin receptor interaction region). Both PIR and SH2 domains interact with the insulin receptor, but their relative role varies considering the member of the Grb7 family and the tyrosine kinase receptor. In the case of Grb14, PIR is the main binding domain and is sufficient to inhibit the insulin receptor kinase activity. We have proposed, on the basis of NMR measurements, that PIR lacks ordered structure and presents a high flexibility, although remaining fully active. To complement this first study, we have used small-angle x-ray scattering in solution together with a modeling approach representing the PIR domain as a chain of pseudo residues. Circular dichroism experiments were also performed in the presence of variable amounts of trifluoroethanol. These observations, together with an ensemble of sequence analyses and previous NMR results, all support the view of PIR as essentially unstructured but with a potentially structured short stretch encompassing residues 399-407. This stretch, which may be only structured transiently in the isolated molecule, could play a major role in Grb14 PIR binding to a biological partner by undergoing a structural transition.
Laboratoire de Cristallographie et RMN Biologiques, CNRS UMR 8015, Faculté de Pharmacie, Université Paris 5, 75270 Paris Cédex 06, France.
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Moncoq
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Broutin
C T
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Craescu
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P
Vachette
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Ducruix
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Durand
D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D003198:Computer Simulation; D008956:Models, Chemical; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011487:Protein Conformation; D011489:Protein Denaturation; D017510:Protein Folding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D011506:Proteins / Q000032:analysis* / Q000737:chemistry*; D011972:Receptor, Insulin / Q000737:chemistry*; D020539:Sequence Analysis, Protein / Q000379:methods*; D013329:Structure-Activity Relationship; D014961:X-Ray Diffraction / Q000379:methods*
D048868:Adaptor Proteins, Signal Transducing; C490976:Grb14 protein, rat; D011506:Proteins; D011972:Receptor, Insulin