An intramolecular interaction between SH2-kinase linker and kinase domain is essential for the catalytic activity of protein-tyrosine kinase-6.
Protein-tyrosine kinase-6 (PTK6, also known as Brk) is a non-receptor tyrosine kinase that contains SH3, SH2, and catalytic (Kinase) domains. We have identified an intramolecular interaction between the linker (Linker) region connecting the SH2 and Kinase domains and the Kinase domain. Residue Trp-184 within the Linker region is essential for the Linker-Kinase interaction but not for the Linker-SH3 interaction. A recombinant PTK6 Kinase domain connected to the Linker region had catalytic activity in terms of autophosphorylation, phosphorylation of a PTK6 substrate, BKS, and phosphorylation of an oligopeptide substrate, whereas the Kinase domain itself, or one connected to a Linker region containing a W184A substitution, did not. The introduction of the W184A mutation into PTK6 also abrogated autophosphorylation and phosphorylation of another PTK6 substrate, Sam68, as well as phosphorylation of intracellular proteins. It also abolished the ability of PTK6 to promote proliferation and prevent apoptosis of HEK 293 cells, as well as to permit anchorage-independent colony formation. Therefore, unlike Src family members, in which the Linker-Kinase interaction inhibits catalytic activity, in PTK6 this interaction has an essential positive role.
Kim
Han Ie
HIe
National Research Laboratory of Cellular Biochemistry, Department of Biochemistry, College of Science, and Protein Network Research Center, Yonsei University, Seoul 120-749, Korea.
Lee
Seung-Taek
ST
D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D017209:Apoptosis; D015153:Blotting, Western; D002384:Catalysis; D002460:Cell Line; D049109:Cell Proliferation; D002470:Cell Survival; D004268:DNA-Binding Proteins; D004926:Escherichia coli / Q000378:metabolism; D005982:Glutathione Transferase / Q000378:metabolism; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology; D047468:Immunoprecipitation; D007700:Kinetics; D008969:Molecular Sequence Data; D009154:Mutation; D009363:Neoplasm Proteins / Q000737:chemistry / Q000378:metabolism*; D010455:Peptides / Q000737:chemistry; D010766:Phosphorylation; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011505:Protein-Tyrosine Kinases / Q000737:chemistry / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry; D011994:Recombinant Proteins / Q000737:chemistry; D017386:Sequence Homology, Amino Acid; D020349:Surface Plasmon Resonance; D013997:Time Factors; D014162:Transfection; D014364:Tryptophan / Q000737:chemistry; D018909:src Homology Domains
D048868:Adaptor Proteins, Signal Transducing; D004268:DNA-Binding Proteins; C489559:KHDRBS1 protein, human; D009363:Neoplasm Proteins; D010455:Peptides; D016601:RNA-Binding Proteins; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D014364:Tryptophan; D006861:Hydrogen Peroxide; D005982:Glutathione Transferase; D011505:Protein-Tyrosine Kinases; C088375:PTK6 protein, human