Although protein phosphorylation sites can be reliably identified with high-resolution mass spectrometry, the experimental approach is time-consuming and resource-dependent. Furthermore, it is unlikely that an experimental approach could catalog an entire phosphoproteome. Computational prediction of phosphorylation sites provides an efficient and flexible way to reveal potential phosphorylation sites and provide hypotheses in experimental design. Musite is a tool that we previously developed to predict phosphorylation sites based solely on protein sequence. However, it was not comprehensively applied to plants. In this study, the phosphorylation data from Arabidopsis thaliana, B. napus, G. max, M. truncatula, O. sativa, and Z. mays were collected for cross-species testing and the overall plant-specific prediction as well. The results show that the model for A. thaliana can be extended to other organisms, and the overall plant model from Musite outperforms the current plant-specific prediction tools, Plantphos, and PhosphAt, in prediction accuracy. Furthermore, a comparative study of predicted phosphorylation sites across orthologs among different plants was conducted to reveal potential evolutionary features. A bipolar distribution of isolated, non-conserved phosphorylation sites, and highly conserved ones in terms of the amino acid type was observed. It also shows that predicted phosphorylation sites conserved within orthologs do not necessarily share more sequence similarity in the flanking regions than the background, but they often inherit protein disorder, a property that does not necessitate high sequence conservation. Our analysis also suggests that the phosphorylation frequencies among serine, threonine, and tyrosine correlate with their relative proportion in disordered regions. Musite can be used as a web server (http://musite.net) or downloaded as an open-source standalone tool (http://musite.sourceforge.net/).
Keywords: Musite; machine-learning; orthologous; phosphorylation; plant evolution; protein disorder.