Side-chain dynamics of the SAP SH2 domain correlate with a binding hot spot and a region with conformational plasticity.
X-linked lymphoproliferative disease is caused by mutations in the protein SAP, which consists almost entirely of a single SH2 domain. SAP interacts with the Tyr281 site of the T<-->B cell signaling protein SLAM via its SH2 domain. Interestingly, binding is not dependent on phosphorylation but does involve interactions with residues N-terminal to the Tyr. We have used 15N and 2H NMR relaxation experiments to investigate the motional properties of the SAP SH2 domain backbone amides and side-chain methyl groups in the free protein and complexes with phosphorylated and non-phosphorylated peptides derived from the Tyr281 site of SLAM. The most mobile methyl groups are in side-chains with large RMSD values between the three crystal structures of SAP, suggesting that fast time-scale dynamics in side-chains is associated with conformational plasticity. The backbone amides of two residues which interact with the C-terminal part of the peptides experience fast time-scale motions in the free SH2 domain that are quenched upon binding of either the phosphorylated or non-phosphorylated peptide. Of most importance, the mobility of methyl groups in and around the binding site for residues in the N-terminus of the peptide is significantly restricted in the complexes, underscoring the dominance of this interaction with SAP and demonstrating a correlation between changes in rapid side-chain motion upon binding with local binding energy.
Finerty
Patrick J
PJ
Structural Biology and Biochemistry, The Hospital for Sick Children, 555 University Avenue, M5G 1X8, Toronto, Ont., Canada.
Muhandiram
Ranjith
R
Forman-Kay
Julie D
JD
D001665:Binding Sites; D002352:Carrier Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D007700:Kinetics; D008024:Ligands; D019563:Mechanics; D008958:Models, Molecular; D019906:Nuclear Magnetic Resonance, Biomolecular; D010446:Peptide Fragments / Q000737:chemistry* / Q000378:metabolism; D019000:Phosphotyrosine / Q000378:metabolism; D011485:Protein Binding; D017510:Protein Folding; D000071179:Signaling Lymphocytic Activation Molecule Associated Protein; D013816:Thermodynamics; D014443:Tyrosine / Q000378:metabolism; D018909:src Homology Domains
D002352:Carrier Proteins; D047908:Intracellular Signaling Peptides and Proteins; D008024:Ligands; D010446:Peptide Fragments; C114953:SH2D1A protein, human; D000071179:Signaling Lymphocytic Activation Molecule Associated Protein; D019000:Phosphotyrosine; D014443:Tyrosine