Figure 2

Bioinformatic analyses of potential Chk1 substrates based on phospho-peptides identified by mass spectrometry. (a) Schematic for in vitro labeling and identification of Chk1 substrates. (b) as-Chk1 uses N6B-ATPγS as detected by antibodies recognizing thio-phosphorylation (thio-phosphate ester (TPE) moiety). (c) Euler diagram depicting the proportion of phospho-sites identified known to occur in vivo [11, 12]. (d) Classification of identified Chk1 substrates based on biological processes (Gene Ontology Consortium). Proteins involved in nucleic acid metabolism were further classified. (e) Euler diagram depicting the proportion of proteins found in this screen with links with the DNA-damage response (DDR; comparison with [13, 14, 21, 23–25]). (f) Frequencies of amino acids surrounding phospho-sites identified in our screen. The x-axis represents the sequence window, with the phosphorylated residue in the middle. Amino acid size depicts fold enrichment (positive, above y-axis) or under-representation (negative, below y-axis) after normalization to amino acid occurrences in the human proteome. Amino acid colors: black, hydrophobic; blue, basic; red, acidic; green, polar; purple, ester. Residues shown in pink were never found in a given position. Note that phospho-peptides containing cysteine were not recovered due to methodological limitations [16]. Diagrams were made with IceLogo software [61]. (g) IceLogo for phospho-peptides with R/K at -3.