Comparative Ser/Thr/Tyr phosphoproteomics between two mycobacterial species: the fast growing Mycobacterium smegmatis and the slow growing Mycobacterium bovis BCG

Abstract

Ser/Thr/Tyr protein phosphorylation plays a critical role in regulating mycobacterial growth and development. Understanding the mechanistic link between protein phosphorylation signaling network and mycobacterial growth rate requires a global view of the phosphorylation events taking place at a given time under defined conditions. In the present study we employed a phosphopeptide enrichment and high throughput mass spectrometry-based strategy to investigate and qualitatively compare the phosphoproteome of two mycobacterial model organisms: the fast growing Mycobacterium smegmatis and the slow growing Mycobacterium bovis BCG. Cells were harvested during exponential phase and our analysis detected a total of 185 phospho-sites in M. smegmatis, of which 106 were confidently localized [localization probability (LP) = 0.75; PEP = 0.01]. By contrast, in M. bovis BCG the phosphoproteome comprised 442 phospho-sites, of which 289 were confidently localized. The percentage distribution of Ser/Thr/Tyr phosphorylation was 39.47, 57.02, and 3.51% for M. smegmatis and 35, 61.6, and 3.1% for M. bovis BCG. Moreover, our study identified a number of conserved Ser/Thr phosphorylated sites and conserved Tyr phosphorylated sites across different mycobacterial species. Overall a qualitative comparison of the fast and slow growing mycobacteria suggests that the phosphoproteome of M. smegmatis is a simpler version of that of M. bovis BCG. In particular, M. bovis BCG exponential cells exhibited a much more complex and sophisticated protein phosphorylation network regulating important cellular cycle events such as cell wall biosynthesis, elongation, cell division including immediately response to stress. The differences in the two phosphoproteomes are discussed in light of different mycobacterial growth rates.

The transpeptidase PbpA and noncanonical transglycosylase RodA of Mycobacterium tuberculosis play important roles in regulating bacterial cell lengths

“…8). Our results are in agreement with an independent study wherein RodA was identified to be phosphorylated at Thr-463 in Mycobacterium bovis BCG (71). In vitro kinase assays with purified STPKs showed that PknB and PknH are the most likely STPKs to mediate phosphorylation of RodA (Fig.…”

CTL0511 from Chlamydia trachomatis Is a Type 2C Protein Phosphatase with Broad Substrate Specificity

“…rCTL0511 was incubated with okadaic acid (OA) (17), cyclosporine (CsA), or sodium pyrophosphate (SPP), and phosphatase activity was assayed at 37°C with 25 mM MnCl 2 and pNPP as a substrate. Relative activity was calculated as the percentage of activity remaining in the presence of the inhibitor versus the activity in assay buffer containing DMSO.…”

“…Phosphorylation of protein substrates can lead to direct modulation of protein function or alteration of gene expression on a global level, in the case of activation of transcription factors or signaling pathways (13). In bacteria, Ser, Thr, and Tyr protein phosphorylation, which was once considered to be exclusive to eukaryotes, has gained increasing attention due to its role in major cellular events, including regulation of virulence, stress responses, and differentiation (11,(14)(15)(16)(17).…”

Targeted treatment for bacterial infections: prospects for pathogen-specific antibiotics coupled with rapid diagnostics

“…43 Repurposing drugs designed for other indications can also produce new antibiotics or anti-virulence agents, with the added benefit of known safety and pharmacokinetic profiles. 44,45 …”