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Phosphorylation site specificity of the Pak‐mediated regulation of Raf‐1 and cooperativity with Src
Author(s) -
King Alastair J.,
Wireman Randall S.,
Hamilton Mark,
Marshall Mark S.
Publication year - 2001
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/s0014-5793(01)02425-5
Subject(s) - phosphorylation , serine , phosphorylation cascade , proto oncogene tyrosine protein kinase src , protein phosphorylation , tyrosine phosphorylation , biochemistry , cooperativity , tyrosine , kinase , phosphoserine , chemistry , dephosphorylation , microbiology and biotechnology , peptide , biology , protein kinase a , phosphatase
The p21‐activated kinase, Pak, has recently been shown to phosphorylate Raf‐1 on serine 338 (S338), a critical regulatory residue. The specificity requirements for Pak‐mediated phosphorylation of S338 were examined by substitution analysis of Raf‐1 peptides and conserved region 3 (CR3) proteins. Phosphorylation was found to be very sensitive to alterations in amino acid side chains proximal to S338. Loss of N‐terminal arginines resulted in decreased peptide phosphorylation while loss of these residues, as well as C‐terminal glutamates and bulky C‐terminal hydrophobic residues, decreased phosphorylation of the CR3 protein. Phosphorylation of Raf‐1 on tyrosine 341 is significant in epidermal growth factor‐ and Src‐mediated signaling, suggesting that cooperativity may exist between Pak and Src phosphorylation of Raf‐1. Purified Pak and Src were found not to be cooperative in phosphorylating peptides or purified CR3 protein. However, the phosphorylation of Raf‐1 S338 by Pak was increased in the presence of Src. The complexity of this signaling module could thus account for the different levels of Raf‐1 activation required for fulfillment of different biological roles within the cell.