Systematic Analysis of Protein Phosphorylation Networks From Phosphoproteomic Data

Abstract

In eukaryotes, hundreds of protein kinases (PKs) specifically and precisely modify thousands of substrates at specific amino acid residues to faithfully orchestrate numerous biological processes, and reversibly determine the cellular dynamics and plasticity. Although over 100,000 phosphorylation sites (p-sites) have been experimentally identified from phosphoproteomic studies, the regulatory PKs for most of these sites still remain to be characterized. Here, we present a novel software package of iGPS for the prediction of in vivo site-specific kinase-substrate relations mainly from the phosphoproteomic data. By critical evaluations and comparisons, the performance of iGPS is satisfying and better than other existed tools. Based on the prediction results, we modeled protein phosphorylation networks and observed that the eukaryotic phospho-regulation is poorly conserved at the site and substrate levels. With an integrative procedure, we conducted a large-scale phosphorylation analysis of human liver and experimentally identified 9719 psites in 2998 proteins. Using iGPS, we predicted a human liver protein phosphorylation networks containing 12,819 potential site-specific kinase-substrate relations among 350 PKs and 962 substrates for 2633 p-sites. Further statistical analysis and comparison revealed that 127 PKs significantly modify more or fewer p-sites in the liver protein phosphorylation networks against the whole human protein phosphorylation network. The largest data set of the human liver phosphoproteome together with computational analyses can be useful for further experimental consideration. This work contributes to the understanding of phosphorylation mechanisms at the systemic level, and provides a powerful methodology for the general analysis of in vivo post-translational modifications regulating sub-proteomes. Molecular & Cellular

Analysis of the Candida albicans Phosphoproteome

“…2B). This distribution is similar to what has been observed in other eukaryotic organisms, including C. neoformans (S, 81.7%; T, 17%; and Y, 1.3%) (24), Tetrahymena thermophila (S, 80.5%; T, 15.7%; and Y, 3.8%) (53), and humans (S, 85.3%; T, 12.9%; and Y, 1.8%) (54). Phosphorylation can also occur on aspartate, lysine, arginine, and histidine residues; however, those phosphorylations are comparatively difficult to detect under our experimental conditions due to their labile character under acidic conditions (55,56).…”

Robust phosphoproteome enrichment using monodisperse microsphere–based immobilized titanium (IV) ion affinity chromatography

“…In turn, fractionation or multidimensional separations combined with Ti 4 + -IMAC are needed for resolving the complexity of cellular phosphoproteome. The Ti 4 + -IMAC method can also be combined with multidimensional chromatographic strategies 41,56,82 . To further exemplify the performance of Ti 4 + -IMAC for large-scale phosphoproteomic analysis, we combined the well-established low-pH SCX chromatography 12,54 with Ti 4 + -IMAC for the analysis of dimethyl-labeled MCF-7 samples.…”

Convergent signaling pathways—interaction between methionine oxidation and serine/threonine/tyrosine O-phosphorylation

“…However, the common occurrence of Met adjacent to Ser/ Thr/Tyr residues (Rao et al 2013) and the frequent occurrence of MetSO (Ghesquière et al 2011;Liu et al 2013;Salvato et al 2014) near O-phosphorylation-sites (Song et al 2012) suggests that crosstalk between these two PTM's is widespread. (Hardin et al 2009;Miernyk et al 2009), oxidation of Met might directly inhibit O-phosphorylation of a nearby Ser/Thr/Tyr-residue by interfering with the kinase recognition/binding (Fig.…”

“…It is notable that mitochondria are the major site of cellular ROS production (Møller 2001;Brand 2010). Song et al (2012) reported hundreds of oxidized Met residues within thousands of phosphopeptides identified from human liver.…”