Beimling P, Niehof M, Radziwill G, Moelling K
Max-Planck-Institut fuer Molekulare Genetik, Abt. Schuster, Berlin, Dahlem, FRG.
Biochem Biophys Res Commun. 1994 Oct 28;204(2):841-8. doi: 10.1006/bbrc.1994.2536.
The kinase negative aminoterminal domain of c-Raf-1 expressed as glutathione S-transferase fusion protein was phosphorylated in vitro after treatment with lysates from A431 cells and subsequent in vitro protein kinase assay. This phosphorylation was independent of stimulation of the cells with EGF; it occurred exclusively on serine and was mapped to Ser259. The identical site of c-Raf-1 was phosphorylated in A431 cells by metabolic labelling in vivo. The kinase binding domain was mapped by various GST-Raf deletion mutants to c-Raf-1 aminoacid residues 181 to 255.
作为谷胱甘肽S-转移酶融合蛋白表达的c-Raf-1激酶阴性氨基末端结构域，在用A431细胞裂解物处理并随后进行体外蛋白激酶测定后，在体外被磷酸化。这种磷酸化与用表皮生长因子（EGF）刺激细胞无关；它仅发生在丝氨酸上，并定位到Ser259。通过体内代谢标记，c-Raf-1的相同位点在A431细胞中被磷酸化。通过各种GST-Raf缺失突变体将激酶结合结构域定位到c-Raf-1的181至255位氨基酸残基。
