Whole proteome analysis of post-translational modifications: Applications of mass-spectrometry for proteogenomic annotation

Abstract

While bacterial genome annotations have significantly improved in recent years, techniques for bacterial proteome annotation (including post-translational chemical modifications, signal peptides, proteolytic events, etc.) are still in their infancy. At the same time, the number of sequenced bacterial genomes is rising sharply, far outpacing our ability to validate the predicted genes, let alone annotate bacterial proteomes. In this study, we use tandem mass spectrometry (MS/MS) to annotate the proteome of Shewanella oneidensis MR-1, an important microbe for bioremediation. In particular, we provide the first comprehensive map of post-translational modifications in a bacterial genome, including a large number of chemical modifications, signal peptide cleavages, and cleavages of N-terminal methionine residues. We also detect multiple genes that were missed or assigned incorrect start positions by gene prediction programs, and suggest corrections to improve the gene annotation. This study demonstrates that complementing every genome sequencing project by an MS/MS project would significantly improve both genome and proteome annotations for a reasonable cost.

Augmented Annotation of the Schizosaccharomyces pombe Genome Reveals Additional Genes Required for Growth and Viability

“…A number of pipelines have successfully exploited high-throughput tandem mass-spectrometry (MS/MS) data to predict novel proteins in H. sapiens (Fermin et al 2006;Tanner et al 2007;Bitton et al 2010), Caenorhabditis elegans (Schrimpf et al 2009), Drosophila melanogaster (Schrimpf et al 2009), Arabidopsis thaliana (Castellana et al 2008), and bacteria (Gupta et al 2007). We have therefore applied a pipeline that integrates this proteogenomics approach (Bitton et al 2010) with comparative genomics and genomewide domain prediction to augment the current annotation of S. pombe.…”

Post-Translational Modifications of Desulfovibrio vulgaris Hildenborough Sulfate Reduction Pathway Proteins

“…Using these datasets, 24 Mascot MS/MS Ion searches, including decoy database searches, were conducted to 25 identify potential PTMs in D. vulgaris. Recently, a number of similar studies have been 26 reported in other organisms [12][13][14] . Many PTMs are known to play critical roles in bacterial 27 regulation.…”

“…In this search, a total of 6220 unique peptide 13 sequences passed the filtering thresholds. Of these, 309 (~5%) were identified as having 14 an unlabeled peptide N-terminus. A total of 2665 sequences contained one or more 15 lysines, and of these, only 14 (~0.5%) were identified as having unlabeled K. The overall 16 trend observed is that iTRAQ labeling is highly efficient for K side chains, but peptide N-17 termini are slightly less reactive.…”

Proteomic analysis of post translational modifications in cyanobacteria

“…Moreover, the same experimental proteomic datasets are also useful in identifying PTMs on a proteome wide level [100,101]. Our group developed a proteogenomic approach to carry out genome annotation and whole-proteome analysis of PTMs in prokaryotes by employing high resolution and high accuracy MS data, and we used this approach to study the global PTMs in the model cyanobacterium Synechococcus 7002 [27].…”