Probing the communication between the regulatory and catalytic domains of a protein tyrosine kinase, Csk.
Protein tyrosine kinases (PTKs) are important regulators of mammalian cell function and their own activities are tightly regulated. Underlying their tight regulation, all PTKs contain multiple regulatory domains in addition to a catalytic domain. C-terminal Src kinase (Csk) contains a catalytic domain and a regulatory region, consisting of an SH3 and an SH2 domain. In this study, we probed the communication between the regulatory and catalytic domains of Csk. First, kinetic characterization of SH3 and SH2 domain deletion mutants demonstrated that the SH3 and SH2 domains were crucial in maintaining the full activity of Csk, but were not directly involved in Csk recognition of its physiological substrate, Src. Second, highly conserved Trp188, corresponding to a key residue in domain-domain communication in other PTKs, was found to be important for maintaining the active structure of Csk by the presence of the regulatory region, but not required for Csk activation triggered by a phosphopeptide binding to the SH2 domain. Third, structural alignment indicated that the presence of the regulatory domains modulated the conformation of multiple substructures in the catalytic domain, some directly and others remotely. Mutagenic and kinetic studies supported this assignment. This report extended previous studies of Csk domain-domain communication, and provided a foundation for further detailed investigation of this communication.
Lin
Xiaofeng
X
Department of Cell and Molecular Biology, University of Rhode Island, Kingston, Rhode Island 02881, USA.
Ayrapetov
Marina K
MK
Lee
Sungsoo
S
Parang
Keykavous
K
Sun
Gongqin
G
D000409:Alanine / Q000235:genetics; D000818:Animals; D000081247:CSK Tyrosine-Protein Kinase; D020134:Catalytic Domain / Q000235:genetics; D002645:Chickens; D018360:Crystallography, X-Ray; D004789:Enzyme Activation / Q000235:genetics; D010649:Phenylalanine / Q000235:genetics; D010748:Phosphopeptides / Q000737:chemistry / Q000378:metabolism; D019000:Phosphotyrosine / Q000737:chemistry / Q000378:metabolism; D017354:Point Mutation; D011487:Protein Conformation; D011505:Protein-Tyrosine Kinases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D016392:Proto-Oncogene Proteins pp60(c-src) / Q000172:deficiency / Q000235:genetics; D016415:Sequence Alignment; D017384:Sequence Deletion; D013379:Substrate Specificity / Q000235:genetics; D014364:Tryptophan / Q000235:genetics; D018909:src Homology Domains / Q000235:genetics; D019061:src-Family Kinases
D010748:Phosphopeptides; D019000:Phosphotyrosine; D010649:Phenylalanine; D014364:Tryptophan; D011505:Protein-Tyrosine Kinases; D000081247:CSK Tyrosine-Protein Kinase; D016392:Proto-Oncogene Proteins pp60(c-src); D019061:src-Family Kinases; D000409:Alanine