Vesicular Proteome Potential for Vaccine and Biomarker Applications

In pathogenic, Gram-negative bacteria, outer membrane vesicles (OMVs) function by adhering extracellularly to host cells and serving as transportation molecules for immune-modulating compounds.¹ OMVs have been used clinically to enhance immune function when repurposed into antigenic vaccines or biomedical delivery systems.^2,3

The most common source of bacterial food poisoning, Campylobacter jejuni, possesses a vesicular proteome that has not been fully investigated. Because the OMVs almost certainly participate in the pathogenesis and virulence of the food-borne pathogen, a thorough study of these structural components may offer insight into the microbe itself, its function as a pathogen, and its potential application in the production of vaccines.

Recently, Jang et al. (2013) used high-resolution LTQ-Orbitrap mass spectrometry (Thermo Scientific) to explore the OMV proteome derived from C. jejuni clinical strain NCTC11168.⁴Using data derived from electron microscopy, the researchers noted that the desiccated vesicles were spherical, composed of a bilayer, and 50 nm in average diameter. Hydrated vesicles possessed a mean radius of 77.5 ± 4.6 nm, although the researchers indicated that these biologically relevant nano-OMVs may not reflect the full range of OMV size in natural populations. Jang et al. also used gel methodology to examine OMV structure and noted that the OMVs in the sample contained fewer glycoproteins than did whole-cell lysates.

Using high-resolution nano liquid chromatography–tandem mass spectrometry (LC-MS/MS) performed in triplicate, the researchers observed a total of 283 vesicular proteins. They isolated 134 of those proteins with high confidence. They classified these by both subcellular localization and biological function, including 48 membrane proteins (36%), 15 periplasmic proteins (11%), 43 cytosolic proteins (32%) and 28 unknown proteins (21%). Of the unknown proteins, 5 were predicted to have signal peptides and were thus likely periplasmic. The research team also identified 64 enzymes, including 20 oxidoreductases (15%), 18 transferases (13%), 5 hydrolases (4%), 7 lyases (5%), 6 isomerases (5%) and 8 ligases (6%).

Jang et al. compared the 134 strongly identified proteins to 774 genes with known functions identified from whole cell C. jejuni lysates. Notable genetic functions enriched in this study include energy production, respiratory metabolism, flagellar assembly and cell adhesion. The investigators also observed several N-linked glycoproteins within the vesicles.

Using signal peptide predictors for comparison with identified C. jejuni vesicular proteins revealed that 53 proteins (40% total) possess a signal peptide. Of these, 8 vesicular proteins (6% total) likely use the Tat apparatus, and 35 vesicular proteins (26% total) likely localize using the Sec pathway. Searching the identified vesicular proteins for the bacterial N-glycosylation motif revealed that 35 of the proteins contained the motif; only 11 of these could be confirmed.

Overall, Jang et al. assert that C. jejuni OMVs contain a diverse range of proteins, indicating functional roles in several biological processes. They call for further investigation into the specific role of each identified protein, which may lead to the identification of novel antibacterial targets, diagnostic markers and vaccine candidates.

References

1. Kuehn, M.J., and Kesty, N.C. (2005) “Bacterial outer membrane vesicles and the host-pathogen interaction,” Genes and Development, 19 (pp. 2645–55).

2. Collins, B.S. (2011) “Gram-negative outer membrane vesicles in vaccine development,” Discovery Medicine, 12 (pp. 7–15).

3. Kim, J.Y., et al. (2008) “Engineered bacterial outer membrane vesicles with enhanced functionality,” Journal of Molecular Biology, 380 (pp. 51–66).

4. Jang, K.-S., et al. (2013) “Comprehensive proteomic profiling of outer 2 membrane vesicles from Campylobacter jejuni,” Journal of Proteomics, available at http://dx.doi.org/10.1016/j.jprot.2013.12.014.

Post Author: Melissa J. Mayer. Melissa is a freelance writer who specializes in science journalism. She possesses passion for and experience in the fields of proteomics, cellular/molecular biology, microbiology, biochemistry, and immunology. Melissa is also bilingual (Spanish) and holds a teaching certificate with a biology endorsement.