Goals and Challenges in Bacterial Phosphoproteomics

Abstract

Reversible protein phosphorylation at serine, threonine and tyrosine is a well-known dynamic post-translational modification with stunning regulatory and signalling functions in eukaryotes. Shotgun phosphoproteomic analyses revealed that this post-translational modification is dramatically lower in bacteria than in eukaryotes. However, Ser/Thr/Tyr phosphorylation is present in all analysed bacteria (24 eubacteria and 1 archaea). It affects central processes, such as primary and secondary metabolism development, sporulation, pathogenicity, virulence or antibiotic resistance. Twenty-nine phosphoprotein orthologues were systematically identified in bacteria: ribosomal proteins, enzymes from glycolysis and gluconeogenesis, elongation factors, cell division proteins, RNA polymerases, ATP synthases and enzymes from the citrate cycle. While Ser/Thr/Tyr phosphorylation exists in bacteria, there is a consensus that histidine phosphorylation is the most abundant protein phosphorylation in prokaryotes. Unfortunately, histidine shotgun phosphorproteomics is not possible due to the reduced phosphohistidine half-life under the acidic pH conditions used in standard LC-MS/MS analysis. However, considering the fast and continuous advances in LC-MS/MS-based phosphoproteomic methodologies, it is expected that further innovations will allow for the study of His phosphoproteomes and a better coverage of bacterial phosphoproteomes. The characterisation of the biological role of bacterial Ser/Thr/Tyr and His phosphorylations might revolutionise our understanding of prokaryotic physiology.

Automated Phosphopeptide Enrichment for Gram-Positive Bacteria

“…B. subtilis and L. monocytogenes showed the highest percentage of phosphorylation at serine followed by threonine and tyrosine (S/T/Y 67/28/5% for B. subtilis and 56/35/9% for L. monocytogenes ). Our data are in line with previously reported S/T/Y phosphorylation–distribution in B. subtilis and L. monocytogenes , approximately 70/20/10% 2 , 24 , 25 and 65/30/5%, 2 , 20 , 21 respectively. Notably, we observed that threonine was the main site of phosphorylation in S. pyogenes ( Figure 2 A).…”

“…Involvement of protein phosphorylation in the regulation of the central carbon metabolism was proposed in previous studies as a large subset of phosphorylated proteins in bacteria is involved in this pathway. 2 , 5 , 20 , 24 , 27 In line with those findings, gene ontology enrichment analysis revealed enrichment of phosphoproteins involved in carbohydrate metabolism in L. monocytogenes and S. pyogenes . Additionally, we observed enrichment of proteins involved in translation in all three species ( Figure 2 E).…”

“…We observed a S/T/Y phosphorylation distribution of approximately 41/55/4% in S. pyogenes , which is different from the reported distributions in two closely related species, S. pneumoniae and S. thermophilus (approximately 47/44/9% and 43/33/23%, respectively). 2 , 22 , 26 …”

“…In recent years, evidence has accumulated suggesting that S/T/Y phosphorylation is, albeit at a less frequent occurrence, similarly important in prokaryotes. 2 − 4 A large subset of phosphorylated proteins in bacteria is involved in the central carbon metabolism. 5 Furthermore, protein phosphorylation in bacteria mediates different cellular processes required for virulence, including remodeling of the bacterial cell surface, expression of virulence genes, and interference with host signaling pathways.…”

Correction: Yagüe, P., et al. Goals and Challenges in Bacterial Phosphoproteomics. Int. J. Mol. Sci. 2019, 20, 5678

“…The authors wish to make the following corrections to this paper [ 1 ]: The author name “Gemma Fernánez-García” should be “Gemma Fernández-García”. Page 2, first line: “257 proteins” should be “301 proteins”.…”