S79 within the CRIB domain is crucial for autophosphorylation and kinase activity of PAK1.

<p>Overall Structures of CRIP domain of PAK1 and Sequence Comparisons. (<b>A</b>) Ribbon diagram showing a structural overview of CRIP domain of PAK1 (red) with bound Cdc42 (blue). Data Bank ID number 1E0A and visualized using Molsoft ICM Browser (Molsoft, L.L.C., San Diego, CA, USA). (<b>B</b>) Multiple sequence alignment of the amino acid sequence of PAK1 from 17 species. (<b>C</b>) Western blot analysis of phosphorylation of T423, S199, and S144 and of GFP-PAK1^WT or GFP-PAK1^S79A. (<b>D</b>) The kinase activity of PAK1 (GFP-PAK1^WT or GFP-PAK1^S79A) was assessed by <i>in vitro</i> phosphorylation assay using MBP (upper) or DLC1 peptide (lower) as substrate. (<b>E</b>) Western blot analysis of the effect of S79A mutation on the PAK1 autophosphorylation using anti-T423 and anti-S144 phospho-specific PAK1 antibodies. Cells were unstimulated (−) or stimulated (+) by EGF (100 ng/ml).