Recombinant Csk expressed in Escherichia coli is autophosphorylated on tyrosine residue(s).
The C-terminal src kinase (Csk) is responsible for the phosphorylation of the carboxy-terminal tyrosine of several tyrosine kinases of the Src family. This phosphorylation site has a negative regulatory function. Csk is unique among the members of the protein tyrosine kinase family because it lacks the conserved tyrosine autophosphorylation site and has been thought to be devoid of autophosphorylation activity. Using the glutathione S-transferase (GST) bacterial expression system, we have produced large amounts of a chimeric rat Csk protein. We have determined that the GST-Csk fusion protein isolated from bacteria is autophosphorylated on tyrosine residue(s). GST-Csk and purified Csk are capable of undergoing autophosphorylation on tyrosine residue(s) in vitro. The GST-Csk fusion protein also phosphorylates exogenous substrates, including the heteropolymer poly-Glu/Tyr and enolase. This is the first indication that Csk is autophosphorylated on tyrosine residue(s) both in vivo in bacteria expressing Csk cDNA and in vitro. These findings suggest that the autophosphorylation of Csk might play a role in the regulation of its kinase activity as well as its binding to other cellular proteins.
Bougeret
C
C
Institut Cochin de Génétique Moléculaire, U332 INSERM, Université Paris V, France.
Rothhut
B
B
Jullien
P
P
Fischer
S
S
Benarous
R
R
D000911:Antibodies, Monoclonal / Q000276:immunology; D000081247:CSK Tyrosine-Protein Kinase; D004926:Escherichia coli / Q000378:metabolism*; D005982:Glutathione Transferase / Q000378:metabolism; D010766:Phosphorylation; D011505:Protein-Tyrosine Kinases / Q000276:immunology / Q000378:metabolism*; D016392:Proto-Oncogene Proteins pp60(c-src) / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism*; D014443:Tyrosine / Q000378:metabolism*; D019061:src-Family Kinases
D000911:Antibodies, Monoclonal; D011993:Recombinant Fusion Proteins; D014443:Tyrosine; D005982:Glutathione Transferase; D011505:Protein-Tyrosine Kinases; D000081247:CSK Tyrosine-Protein Kinase; D016392:Proto-Oncogene Proteins pp60(c-src); D019061:src-Family Kinases