title: Study of O-Phosphorylation Sites in Proteins Involved in Photosynthesis-Related Processes in Synechocystis sp. Strain PCC 6803: Application of the SRM Approach.
abstract: O-Phosphorylation has been shown in photosynthesis-related proteins in a cyanobacterium Synechocystis sp. strain PCC 6803 (thereafter Synechocystis 6803), suggesting that phosphorylation of S, T, and Y residues might be important in photosynthesis-related processes. Investigation of biological roles of these phosphorylation events requires confident knowledge of the phosphorylated sites and prospects for their individual assessment. We performed phosphoproteomic analysis of Synechocystis 6803 using TiO2enrichment of the phosphopeptides, followed by LC-MS/MS, and discovered 367 phosphorylation sites in 190 proteins participating in various cellular functions. Furthermore, we focused on the large group of phosphoproteins that are involved in light harvesting, photosynthesis-driven electron flow, photoprotection, and CO2fixation. The SRM approach was applied to verify/improve assignments of phosphorylation sites in these proteins and to investigate possibilities for analysis of phosphopeptide isomers. The SRM assays were designed for peptides comprising 45 phosphorylation sites. The assays contain peptide iRT values and Q1/Q3 transitions comprising those discriminating between phosphopeptide isoforms. The majority of investigated phosphopeptides and phosphorylated isoforms could be individually assessed with the SRM technique. The assays could be potentially used in future quantitative studies to evaluate an extent of phosphorylation in photosynthesis-related proteins in Synechocystis 6803 cells challenged with various environmental stresses.
authors: Molecular Plant Biology, Department of Biochemistry, University of Turku , FI-20014 Turku, Finland.
authors: Martina
authors: M
authors: Angeleri
authors: Molecular Plant Biology, Department of Biochemistry, University of Turku , FI-20014 Turku, Finland.
authors: Dorota
authors: D
authors: Muth-Pawlak
authors: Molecular Plant Biology, Department of Biochemistry, University of Turku , FI-20014 Turku, Finland.
authors: Eva-Mari
authors: EM
authors: Aro
authors: Molecular Plant Biology, Department of Biochemistry, University of Turku , FI-20014 Turku, Finland.
authors: Natalia
authors: N
authors: Battchikova
mesh_terms: D001426:Bacterial Proteins / Q000378:metabolism* / Q000502:physiology; D001665:Binding Sites; D010748:Phosphopeptides / Q000032:analysis*; D010766:Phosphorylation; D010788:Photosynthesis; D020033:Protein Isoforms; D040901:Proteomics / Q000379:methods; D046939:Synechocystis / Q000737:chemistry*
chemical_list: D001426:Bacterial Proteins; D010748:Phosphopeptides; D020033:Protein Isoforms
keywords: SRM; Synechocystis 6803; cyanobacteria; phosphopeptide isoforms; phosphopeptides; phosphoproteins; phosphoproteomics; photosynthesis; protein phosphorylation; selected reaction monitoring