Submitted to: Applied and Environmental Microbiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 2/5/2006
Publication Date: 4/11/2006
Citation: Donovan, D.M., Dong, S., Garrett, W.M., Rousseau, G.M., Moineau, S., Pritchard, D.G. 2006. Petidoglycan hydrolase fusions maintain parental specificities. Applied and Environmental Microbiology. 72:2988-2996.
Interpretive Summary: Staphylococcus aureus and Streptococcus agalactiae are human pathogens as well as mastitis causing organisms (infections of dairy cattle mammary glands). Badcteria and bacteriophage synthesize endolysins that degrade the bacterial cell wall peptidoglycan. These endolysins can be used as antimicrobials if added exosgenously to the cell. We describe the fusion of endolysin fragments (or modules) to create an antimicrobial that specifically attacks these two unrelated Gram positive pathogens. Avoiding the generalized use of broad range antimicrobials through the use of highly specific peptidoglycan hydrolases (bacterial or bacteriophage endolysins) might help reduce the incidence of antibiotic resistance that plagues therapeutic efforts world wide.
Technical Abstract: The increased incidence of bacterial antibiotic resistance has led to a renewed search for novel antimicrobials. Avoiding the use of broad range antimicrobials through the use of specific peptidoglycan hydrolases (endolysins) might reduce the incidence of antibiotic resistant pathogens worldwide. Staphylococcus aureus and Streptococcus agalactiae are human pathogens and also cause mastitis in dairy cattle. The ultimate goal of this work is to create transgenic cattle that are resistant to mastitis through the expression of antimicrobial protein(s) in their milk. Toward this end, two novel antimicrobials were produced. The (1) full length and (2) 182 amino acid, C-terminal truncated, S. agalactiae bacteriophage B30 endolysin were fused to the mature lysostaphin of Staphylococcus simulans. Both fusions display lytic specificity for streptococcal pathogens and S. aureus. The full lytic ability of the truncated B30 protein also suggests that the SH3b domain at the C-terminus is dispensable. The fusions are active in a milk-like environment. They are also active against some lactic acid bacteria used in cheese and yogurt making but the lytic activities are destroyed during pasteurization (63'C x 30 min.). Immunohistochemical studies indicate that the fusion protein can be expressed in cultured mammalian cells with no obvious deleterious effects on the cells, making it a strong candidate for use in future transgenic mice and cattle. Since the fusion peptidoglycan hydrolase kills multiple human pathogens, it also may prove useful as a highly selective, multi-pathogen targeting antimicrobial agent that could potentially reduce the use of broad range antibiotics in fighting clinical infections.