Proteome analysis of the hyaluronic acid-producing bacterium, Streptococcus zooepidemicus

Abstract

Background: Streptococcus equi subsp. zooepidemicus (S. zooepidemicus) is a commensal of horses and an opportunistic pathogen in many animals and humans. Some strains produce copious amounts of hyaluronic acid, making S. zooepidemicus an important industrial microorganism for the production of this valuable biopolymer used in the pharmaceutical and cosmetic industry. Encapsulation by hyaluronic acid is considered an important virulence factor in other streptococci, though the importance in S. zooepidemicus remains poorly understood. Proteomics may provide a better understanding of virulence factors in S. zooepidemicus, facilitate the design of better diagnostics and treatments, and guide engineering of superior production strains.

Results: Using hyaluronidase to remove the capsule and by optimising cellular lysis, a reference map for S. zooepidemicus was completed. This protocol significantly increased protein recovery, allowing for visualisation of 682 spots and the identification of 86 proteins using mass spectrometry (LC-ESI-MS/MS and MALDI-TOF/TOF); of which 16 were membrane proteins.

Conclusion: The data presented constitute the first reference map for S. zooepidemicus and provide new information on the identity and characteristics of the more abundantly expressed proteins.

Figure 1

2-DE gels of S. zooepidemicus. Proteins were separated using pH gradient 4–7 on 7 cm 12% polyacrylamide gels. Following separation, proteins were stained with silver nitrate. When HA was not removed, poor quality, streaky gels were obtained (A); digestion of HA prior to protein extraction resulted in high quality, streak-free gels (B).

Figure 2

Reference map of S. zooepidemicus (ATCC 35246). Proteins were harvested using hyaluronidase to remove the HA capsule. Proteins were separated using pH gradient 4–7 and 24 cm 12% polyacrylamide gels. Proteins were labelled with cy3 and visualised using a typhoon scanner. Protein spots identified using LC-ESI-MS/MS and MALDI-TOF/TOF numbered and listed in Additional file 1. Numbers not listed in the Additional file did not yield an MS-identified protein hit.

Figure 3

Functional grouping of proteins identified from S. zooepidemicus proteome. Proteins from the glycolysis, pentose phosphate, and HA pathways were identified in the reference map. Biomass constituent enzymes such as amino acid, lipids and peptidoglycan synthesis were also identified. HA: hyaluronic acid, PPP: phosphate pentose pathway; AA: amino acid.