Characterization of Lck-binding elements in the herpesviral regulatory Tip protein.
Herpesvirus saimiri encodes a tyrosine kinase interacting protein (Tip) that binds to T-cell-specific tyrosine kinase Lck via multiple sequence motifs and controls its activity. The regulation of Lck by Tip represents a key mechanism in the transformation of human T-lymphocytes during herpesviral infection. In this study, the interaction of Tip with the regulatory SH3 and SH2 domains of Lck was investigated by biophysical and computational techniques. NMR spectroscopy of isotopically labeled Tip(140-191) revealed that the interaction with the LckSH3 domain is not restricted to the classical proline-rich motif, but also involves the C-terminally adjacent residues which pack into a hydrophobic pocket on the surface of the SH3 domain, thus playing a likely role in mediating binding specificity. Fluorescence binding studies of Tip further demonstrate that Tyr127 in its phosphorylated form represents a strong ligand of the LckSH2 domain, indicating the presence of an additional Lck interaction motif. In contrast, Tyr114, known to be essential for STAT-3 binding, does not interact with the LckSH2 domain, showing that the tyrosines in Tip exhibit distinct binding specificity. The existence of numerous interaction sites between Tip and the regulatory domains of Lck implies a complex regulatory mechanism and may have evolved to allow a gradual regulation of Lck activity in different pathogenic states.
Bauer
Finn
F
Lehrstuhl für Biopolymere, Universität Bayreuth, 95440 Bayreuth, Germany.
Hofinger
Edith
E
Hoffmann
Silke
S
Rösch
Paul
P
Schweimer
Kristian
K
Sticht
Heinrich
H
D000595:Amino Acid Sequence; D002942:Circular Dichroism; D003001:Cloning, Molecular; D017124:Conserved Sequence; D006567:Herpesvirus 2, Saimiriine / Q000737:chemistry* / Q000201:enzymology / Q000378:metabolism*; D057927:Hydrophobic and Hydrophilic Interactions; D007700:Kinetics; D008024:Ligands; D019860:Lymphocyte Specific Protein Tyrosine Kinase p56(lck) / Q000737:chemistry* / Q000378:metabolism*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D019906:Nuclear Magnetic Resonance, Biomolecular; D010750:Phosphoproteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D019000:Phosphotyrosine / Q000737:chemistry; D011485:Protein Binding; D017433:Protein Structure, Secondary; D011522:Protons; D017386:Sequence Homology, Amino Acid; D013050:Spectrometry, Fluorescence; D014764:Viral Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*
D008024:Ligands; D010750:Phosphoproteins; D011522:Protons; D014764:Viral Proteins; C092384:tyrosine kinase interacting protein, Saimiriine herpesvirus 2; D019000:Phosphotyrosine; D019860:Lymphocyte Specific Protein Tyrosine Kinase p56(lck)