Proteogenomic analysis and global discovery of posttranslational modifications in prokaryotes

We describe an integrated workflow for proteogenomic analysis and global profiling of posttranslational modifications (PTMs) in prokaryotes and use the model cyanobacterium Synechococcus sp. PCC 7002 (hereafter Synechococcus 7002) as a test case. We found more than 20 different kinds of PTMs, and a holistic view of PTM events in this organism grown under different conditions was obtained without specific enrichment strategies. Among 3,186 predicted protein-coding genes, 2,938 gene products (>92%) were identified. We also identified 118 previously unidentified proteins and corrected 38 predicted gene-coding regions in the Synechococcus 7002 genome. This systematic analysis not only provides comprehensive information on protein profiles and the diversity of PTMs in Synechococcus 7002 but also provides some insights into photosynthetic pathways in cyanobacteria. The entire proteogenomics pipeline is applicable to any sequenced prokaryotic organism, and we suggest that it should become a standard part of genome annotation projects. Significance Proteogenomics is the application of mass spectrometry-derived proteomic data for testing and refining predicted genetic models. Cyanobacteria, the only prokaryotes capable of oxygenic photosynthesis, are the ancestor of chloroplasts in plants and play crucial roles in global carbon and nitrogen cycles. An integrated proteogenomic workflow was developed, and we tested this system on a model cyanobacterium, Synechococcus 7002, grown under various conditions. We obtained a nearly complete genome translational profile of this model organism. In addition, a holistic view of posttranslational modification (PTM) events is provided using the same dataset, and the results provide insights into photosynthesis. The entire proteogenomics pipeline is applicable to any sequenced prokaryotes and could be applied as a standard part of genome annotation projects.