Surface Localization of Streptococcal Fructose 1,6-bisphosphate Aldolases

Early reports describing the unexpected presence of FBA on the bacterial cell surface resulted from the separation of surface or secreted fractions by gel electrophoresis, and the subsequent identification of proteins by mass spectroscopy or amino-terminal amino acid sequencing. Using these approaches, FBA (often with other cytosolic proteins) was initially identified as a surface-associated or - secreted protein in several species of streptococci, including S. pyogenes (Lei et al. 2000), S. oralis (Wilkins et al. 2003), S. pneumoniae (Ling et al. 2004), S. agalac- tiae (Fluegge et al. 2004), and, more recently, S. suis (Wu et al. 2008).

S. pyogenes (or Group A streptococcus) causes human infections including pharyngitis, impetigo, necrotizing fasciitis, and streptococcal toxic shock syndrome (Walker et al. 2014). Lei et al. prepared concentrated culture supernatants from representative serotype M1 and M3 strains and identified 44 distinct proteins, which were present in both strains, by two-dimensional (2D) gel electrophoresis and amino-terminal amino acid sequencing (Lei et al. 2000). Eight of the identified proteins, including FBA, were glycolytic enzymes and several of these were present in high abundance in mid-log-phase culture supernatants, suggesting specific and active secretion rather than just passive release (Lei et al. 2000).

S. oralis is a member of the mitis group of oral streptococci. In addition to being a component of normal dental plaque, it is also associated with extra-oral diseases including endocarditis. Wilkins et al. identified 27 proteins, including FBA, from zwittergent-extracted surface preparations of S. oralis strain 176 N which had been separated using 2D gel electrophoresis. No recognizable secretion signals anchoring LPXTG- or choline-binding motifs could be identified in these proteins (Wilkins et al. 2003). In this study, FBA was found in multiple forms with different isoelectric points, possibly indicating the presence of posttranslational modifications (Wilkins et al. 2003).

S. agalactiae (or Group B streptococcus) is a leading cause of sepsis and meningitis in neonates (Landwehr-Kenzel and Henneke 2014; see also Chapter 10). To identify proteins released from S. agalactiae, supernatants from stationary and logarithmic cultures were concentrated and separated by SDS- PAGE (Fluegge et al. 2004). Subsequent amino-terminal amino acid sequencing of proteins identified FBA among the secreted proteins, many of which were immunogenic as they were recognized in immunoblots by sera from infected neonates or healthy adults (Fluegge et al. 2004). Similarly in S. suis serotype 9, immunoblotting of extracted cell-wall proteins with immune sera identified FBA as one of eight cell-wall-associated immunogenic proteins (Wu et al. 2008).

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