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Analysis of pp60 c‐src tyrosine kinase activity and phosphotyrosyl phosphatase activity in human colon carcinoma and normal human colon mucosal cells
Author(s) -
DeSeau Virginia,
Rosen Neal,
Bolen Joseph B.
Publication year - 1987
Publication title -
journal of cellular biochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.028
H-Index - 165
eISSN - 1097-4644
pISSN - 0730-2312
DOI - 10.1002/jcb.240350205
Subject(s) - proto oncogene tyrosine protein kinase src , sodium orthovanadate , microbiology and biotechnology , protein tyrosine phosphatase , phosphatase , in vitro , chemistry , biology , tyrosine kinase , tyrosine , kinase , biochemistry , phosphorylation , signal transduction
We have compared the level of phosphotyrosyl phosphatase activity in lysates from normal human colon mucosal cells and human colon carcinoma cells and analyzed the effect of incubating these cells with sodium orthovanadate, an inhibitor of phosphotyrosyl phosphatase activity, on the relative abundance of acid‐stable phosphotyrosine and on in vitro protein kinase activity of pp60 c‐src . Additionally, we compared the effect of lysing these cells in buffer containing only nonionic detergents with RIPA buffer, which contains both sodium dodecyl sulfate and deoxycholate, on the in vitro kinase activity of pp60 c‐src . Our results show that the level of detectable phosphotyrosyl phosphatase activity in lysates derived from normal colon cells and colon carcinoma cells is very similar. Additionally, the abundance of acid‐stable phosphotyrosine in these cells cultured in the absence or presence of vanadate is not significantly different. However, incubation of these cells with vanadate significantly stimulates the activity of pp60 c‐src derived from the normal colon cells in immune‐complex kinase assays, while having no detectable effect on the activity of pp60 c‐src from the colon tumor cells. The in vitro protein kinase activity of pp60 c‐src derived from RIPA buffer lysates of colon carcinoma cells was found to be elevated five‐ to sevenfold when compared with pp60 c‐src from these same cells lysed in buffer containing only Nonidet‐P 40 as a detergent. The type of lysis buffer did not effect the activity of pp60 c‐src from normal colon mucosal cells. These results provide additional evidence that the activity of pp60 c‐src may be regulated differently in colon carcinoma and normal colon mucosal cells.

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