Activation of the Src-family tyrosine kinase Hck by SH3 domain displacement.
The protein Hck is a member of the Src family of non-receptor tyrosine kinases which is preferentially expressed in haematopoietic cells of the myeloid and B-lymphoid lineages. Src kinases are inhibited by tyrosine-phosphorylation at a carboxy-terminal site. The SH2 domains of these enzymes play an essential role in this regulation by binding to the tyrosine-phosphorylated tail. The crystal structure of the downregulated form of Hck has been determined and reveals that the SH2 domain regulates enzymatic activity indirectly; intramolecular interactions between the SH3 and catalytic domains appear to stabilize an inactive form of the kinase. Here we compare the roles of the SH2 and SH3 domains in modulating the activity of Hck in an investigation of the C-terminally phosphorylated form of the enzyme. We show that addition of the HIV-1 Nef protein, which is a high-affinity ligand for the Hck SH3 domain, to either the downregulated or activated form of Hck causes a large increase in Hck catalytic activity. The intact SH3-binding motif in Nef is crucial for Hck activation. Our results indicate that binding of the Hck SH3 domain by Nef causes a more marked activation of the enzyme than does binding of the SH2 domain, suggesting a new mechanism for regulation of the activity of tyrosine kinases.
Moarefi
I
I
Laboratories of Molecular Biophysics, Howard Hughes Medical Institute, The Rockefeller University, New York 10021, USA.
LaFevre-Bernt
M
M
Sicheri
F
F
Huse
M
M
Lee
C H
CH
Kuriyan
J
J
Miller
W T
WT
D000595:Amino Acid Sequence; D002384:Catalysis; D002460:Cell Line; D015536:Down-Regulation; D004789:Enzyme Activation; D004795:Enzyme Stability; D015693:Gene Products, nef / Q000378:metabolism; D015497:HIV-1 / Q000378:metabolism; D013058:Mass Spectrometry; D008954:Models, Biological; D008969:Molecular Sequence Data; D010766:Phosphorylation; D011485:Protein Binding; D011505:Protein-Tyrosine Kinases / Q000235:genetics / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism*; D051572:Proto-Oncogene Proteins c-hck; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D054311:nef Gene Products, Human Immunodeficiency Virus; D018909:src Homology Domains; D019061:src-Family Kinases / Q000378:metabolism*
D015693:Gene Products, nef; D011518:Proto-Oncogene Proteins; D011994:Recombinant Proteins; D054311:nef Gene Products, Human Immunodeficiency Virus; D011505:Protein-Tyrosine Kinases; D051572:Proto-Oncogene Proteins c-hck; D019061:src-Family Kinases