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Structural elements that regulate pp59c-fyn catalytic activity, transforming potential, and ability to associate with
S H Cheng1, P C Espino, J Marshall
1Laboratory of Cellular Regulation, Genzyme Corporation, Framingham, Massachusetts 01701.
Journal of Virology
|January 1, 1991
Summary
pp59c-fyn tyrosine kinase activity is regulated by phosphorylation sites, similar to other src-family kinases. However, pp59c-fyn
Area of Science:
- Biochemistry
- Molecular Biology
- Oncology
Background:
- pp59c-fyn is a member of the src family of tyrosine kinases.
- It shares high homology with other src-family members, except for its unique amino-terminal region.
- pp59c-fyn is involved in cellular signaling pathways and has been implicated in cancer.
Purpose of the Study:
- To investigate the regulatory mechanisms of pp59c-fyn tyrosine kinase activity.
- To identify key phosphorylation sites and their roles in enzyme activity and cellular transformation.
- To compare the regulatory elements of pp59c-fyn with other src-like tyrosine kinases.
Main Methods:
- Site-directed mutagenesis to alter Tyr-531 and Tyr-420 phosphorylation sites.
- Analysis of enzymatic activity of wild-type and mutant pp59c-fyn proteins.
- Cell transformation assays using NIH 3T3 cells.
- Construction and analysis of pp59c-fyn-pp60c-src chimeras.
Main Results:
- The carboxy terminus of pp59c-fyn is crucial for stable association with middle-T-antigen.
- Tyr-531 and Tyr-420 were identified as in vivo and in vitro phosphorylation sites, respectively.
- Mutation of Tyr-531 to phenylalanine (531F) increased enzymatic activity but less than in pp60c-src.
- The 531F mutant did not induce cell transformation, unlike analogous variants of other src-like kinases.
- Chimeric studies suggested that insufficient enhancement of kinase activity, rather than substrate absence, limits pp59c-fyn's transforming potential.
Conclusions:
- pp59c-fyn possesses regulatory elements analogous to other src-like tyrosine kinases.
- Additional regulatory mechanisms, potentially in the amino-terminal region, repress pp59c-fyn's kinase and transforming activities.
- The limited transforming ability of pp59c-fyn may stem from insufficient kinase activity enhancement, suggesting complex regulation beyond known phosphorylation sites.