Cytolysin A‐mediated protein exportation efficiency and its role in enhancing the fitness of live recombinant Salmonella Typhi vaccine strain

• Published in: Letters in applied microbiology Vol. 74; no. 5; pp. 820 - 830
• Main Authors: Loh, F.‐K., Nathan, S., Chow, S.‐C., Fang, C.‐M.
• Format: Journal Article
• Language: English
• Published: England Oxford University Press 01-05-2022
• Subjects: Antigens; bacterial growth kinetics; Bacterial Proteins - metabolism; ClyA protein; cytolysin A; Cytoplasm; Fitness; Growth rate; Immunogenicity; live vector vaccine; Perforin - genetics; Perforin - metabolism; Periplasmic space; protein exportation; Proteins; Reproductive fitness; Salmonella; Salmonella Typhi; Salmonella typhi - genetics; Translocation; Vaccines; Vaccines, Attenuated; Vaccines, Synthetic - genetics; Vaccines, Synthetic - metabolism; bacterial growth kinetics; Salmonella Typhi; cytolysin A; live vector vaccine; protein exportation
• ISSN: 0266-8254; 1472-765X
• DOI: 10.1111/lam.13669

The genetic fusion of cytolysin A (clyA) to heterologous antigen expressed in live Salmonella vector demonstrated efficient translocation into periplasmic space and extracellular medium. Accumulating evidence has shown that clyA‐mediated antigen delivery improved growth fitness and enhanced immunogenicity of live vector vaccine, but the factors influencing this protein exportation has not been investigated. In this study, Toxoplasma gondii antigen fused at C‐terminal of clyA protein was expressed in live S. Typhi vector via both plasmid and chromosomal‐based expressions. The bivalent strains showed comparable growth rates as monovalent strains, but in varies antigen exportation efficiency. ClyA‐fusion antigen with positive charges was translocated to the extracellular spaces, whereas those with negative charges were retained in the cytoplasm. Furthermore, excessive cellular resources expenditure on antigen expression, especially antigen with larger size, could limit the clyA‐fusion antigen exportation, resulting in undesirable metabolic burden that eventually affects the growth fitness. Altogether, the present work indicates potential linkage of factors mainly on antigen properties and expression platforms that may affect clyA‐mediated antigen delivery to enhance the growth fitness of live vector strain.
Significance and Impact of the Study: Gram‐negative bacteria secrete outer membrane vesicles (OMVs) with immunostimulatory properties and proteoliposome nanostructures, making it highly immunogenic for protein delivery. The fusion of heterologous antigen to cytolysin A (clyA) has proven to direct protein translocation into periplasmic space and secretion into OMVs, thus allowing sufficient antigen expression to stimulate high immunogenicity. This study investigated the effects of clyA‐mediated antigen exportation on the growth fitness of Salmonella Typhi live vector and also uncovered that antigen properties and expression platforms influence the exportation efficiency. The data provide technical insights on the construction of clyA for immunogenic antigen delivery in live vector vaccine.