Fig. 2.

Proteins with highly conserved consensus PKA sites were more likely to be phosphorylated by PKA than proteins with less conserved sites. Shown is a phosphorylation analysis of representative proteins from the following five groups identified by the comparative analysis performed here: (i) proteins with consensus PKA sites conserved to C. albicans (A); (ii) proteins with sites conserved to the sensu lato and petite-negative Saccharomyces species; (iii) proteins with sites conserved amongst the sensu stricto Saccharomyces species (B); (iv) proteins with PKA sites in S. cerevisiae only (C); (v) proteins with no consensus PKA sites (D). For this analysis, the amount of label incorporated into a full-length GST fusion protein from [γ-³²P]ATP was assessed with an in vitro assay performed in the presence or absence of PKA as described in Methods (26, 27). Note that the labeled protein bands have been appropriately lined up to facilitate comparisons between samples. Western immunoblots were performed with an α-GST antibody to quantify the relative amount of fusion protein present. (E) A summary graph indicating the percentage of candidates in each of the above groups that was phosphorylated by PKA.