Title: Engineering Antimicrobials that are Refractory to Resistance Development. Authors
Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: July 10, 2008
Publication Date: July 28, 2009
Citation: Donovan, D.M., Becker, S.C. 2009. Engineering Antimicrobials that are Refractory to Resistance Development. Meeting Abstract. Technical Abstract: Multi-drug resistant pathogens are a serious problem in modern health care and there is a need for novel antimicrobials that are refractory to resistance development. Several US government agencies (FDA, CDC and NIH) recommend avoiding the use of broad range antimicrobials, a practice that is known to support resistance development. Lysostaphin, a peptidoglycan hydrolase (glycyl-glycine endopeptidase) secreted by Staphylococcus simulans to kill S. aureus, has been shown to act with near-species specificity. Although it is a potent antimicrobial, even against antibiotic-resistant strains of S. aureus, plasmid-born factors have emerged that confer resistance to lysostaphin, thereby reducing its antimicrobial usefulness. However, no bacterial strains have yet been identified that resist lysis by their own phage endolysins, making this class of enzymes ideal for antimicrobial development. LysK is a staphylococcal bacteriophage endolysin from phage K that harbors both an amidase and endopeptidase peptidoglycan hydrolase domains. It can lyse many staphylococcal strains, including methicillin-resistant S. aureus (MRSA). We have characterized the LysK cleavage sites and determined that LysK and lysostaphin cleave different bonds in the peptidoglycan of S. aureus. When used in combination, these proteins act synergistically to inhibit the growth of S. aureus, including the MRSA strain USA300. By fusing LysK and lysostaphin we created a chimeric enzyme that retains all three parental peptidoglycan hydrolase activities. These fusions demonstrate an activity in turbidity reduction assays against S. aureus that is greater than either of their parental hydrolases alone. In addition to MRSA, these fusions are also able to lyse the lysostaphin-resistant strains ANG133 (femA::Tn551) and ANG 144 (lyrA::FHX12843) further demonstrating their potential as antimicrobials against even hydrolase-resistant strains. We predict that it will be difficult for any bacterium to simultaneously resist all three lytic activities.