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Table 2.

Phosphorylation sites of human PAK1 detected by mass spectrometry

Site Detected without enrichment (+) or with IMAC Homologs/other family members Putative (roman) and known (italics) kinases
S2   +/IMAC   R, M, H/PAK2, PAK3   
S21*  IMAC   R, M, H/PAK2, PAK3  AUTO/AKT 
S57*  IMAC   R, M, H/PAK2, PAK3  AUTO 
Y131*  +/IMAC   R, M, H/PAK2, PAK3  SRC 
Y142   +/IMAC   R, M, H/PAK2, PAK3   PDGF-R  
Y153   +/IMAC   R, M, H/PAK2, PAK3   
S155/S156   +/IMAC   R, M, H/(S155) PAK 2 only   
S174   +   R, M, H/PAK 3   Casein kinase 1  
T185   +   R, M, H/PAK 2   ERK1/casein kinase 1  
T212*  +/IMAC   R, M, H  ERK1, CDK5, CDC2 
Y285   +   R, M, H/PAK 3   PY-SH2 motif  
Y429   +/IMAC   R, M, H/PAK2, PAK3   
Y464   IMAC   R, M, H/PAK2, PAK3   EGF-R/PY-SH2 motif  
Site Detected without enrichment (+) or with IMAC Homologs/other family members Putative (roman) and known (italics) kinases
S2   +/IMAC   R, M, H/PAK2, PAK3   
S21*  IMAC   R, M, H/PAK2, PAK3  AUTO/AKT 
S57*  IMAC   R, M, H/PAK2, PAK3  AUTO 
Y131*  +/IMAC   R, M, H/PAK2, PAK3  SRC 
Y142   +/IMAC   R, M, H/PAK2, PAK3   PDGF-R  
Y153   +/IMAC   R, M, H/PAK2, PAK3   
S155/S156   +/IMAC   R, M, H/(S155) PAK 2 only   
S174   +   R, M, H/PAK 3   Casein kinase 1  
T185   +   R, M, H/PAK 2   ERK1/casein kinase 1  
T212*  +/IMAC   R, M, H  ERK1, CDK5, CDC2 
Y285   +   R, M, H/PAK 3   PY-SH2 motif  
Y429   +/IMAC   R, M, H/PAK2, PAK3   
Y464   IMAC   R, M, H/PAK2, PAK3   EGF-R/PY-SH2 motif  

Serum-stimulated samples were treated with peroxovanadate and calyculin A to inhibit phosphatases. Numbered phosphorylation sites marked with (*) represents previously confirmed phosphorylation. Phosphorylation sites labeled `IMAC' were only detected after sample enrichment by immobilized metal affinity chromatography. Sites detected without enrichment (C18) are denoted with a `+'. Homologous phosphorylation sites among species (R, rat; M, mouse; H, human) and PAK1 isoforms 1-3 were determined via sequence-comparison analysis using FASTA (http://fasta.bioch.virginia.edu/) (Pearson, 1990). Consensus phosphorylation motifs for putative kinases were analyzed using the Scansite scoring algorithm (http://scansite.mit.edu) (Obenauer et al., 2003). For additional data regarding PAK1 phosphorylation, see the Cell Migration Consortium web site: http://www.cellmigration.org.

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