The Peptide Microarray ChloroPhos1.0: A Screening Tool for the Identification of Arabidopsis thaliana Chloroplast Protein Kinase Substrates.

We designed the peptide microarray ChloroPhos1.0 to screen for substrates of chloroplast protein kinases. The peptides represented on the microarray were selected from phosphoproteomics data, and the identified chloroplast phosphopeptides were spotted as 15-mers on a glass slide with the phosphorylation site centered. Altogether, 905 distinct peptides from chloroplast proteins are present on the array. Here we describe how the array can be used to identify the target protein spectrum of chloroplast kinases. We present the method and discuss limitations and challenges associated with the determination of phosphorylation activity on peptide substrates in vitro.