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Applied Microbiology and Biotechnology

, Volume 99, Issue 15, pp 6315–6326 | Cite as

Discovery of novel S. aureus autolysins and molecular engineering to enhance bacteriolytic activity

  • Daniel C. Osipovitch
  • Sophie Therrien
  • Karl E. Griswold
Biotechnologically relevant enzymes and proteins

Abstract

Staphylococcus aureus is a dangerous bacterial pathogen whose clinical impact has been amplified by the emergence and rapid spread of antibiotic resistance. In the search for more effective therapeutic strategies, great effort has been placed on the study and development of staphylolytic enzymes, which benefit from high potency activity toward drug-resistant strains, and a low inherent susceptibility to emergence of new resistance phenotypes. To date, the majority of therapeutic candidates have derived from either bacteriophage or environmental competitors of S. aureus. Little to no consideration has been given to cis-acting autolysins that represent key elements in the bacterium’s endogenous cell wall maintenance and recycling machinery. In this study, five putative autolysins were cloned from the S. aureus genome, and their activities were evaluated. Four of these novel enzymes, or component domains thereof, demonstrated lytic activity toward live S. aureus cells, but their potencies were 10s to 1000s of times lower than that of the well-characterized therapeutic candidate lysostaphin. We hypothesized that their poor activities were due in part to suboptimal cell wall targeting associated with their native cell wall binding domains, and we sought to enhance their antibacterial potential via chimeragenesis with the peptidoglycan binding domain of lysostaphin. The most potent chimera exhibited a 140-fold increase in lytic rate, bringing it within 8-fold of lysostaphin. While this enzyme was sensitive to certain biologically relevant environmental factors and failed to exhibit a measurable minimal inhibitory concentration, it was able to kill lysostaphin-resistant S. aureus and ultimately proved active in lung surfactant. We conclude that the S. aureus proteome represents a rich and untapped reservoir of novel antibacterial enzymes, and we demonstrate enhanced bacteriolytic activity via improved cell wall targeting of autolysin catalytic domains.

Keywords

MRSA Lysins Lytic enzyme CHAP Lysostaphin Antibiotic 

Notes

Acknowledgments

We would like to thank Dr. Gary Sloan at the University of Alabama for kindly providing the RN4220 strains used in this paper. We would also like to thank Ony, Inc. for supplying the Infasurf used in this study. This work was supported in part by R21 grant 1R21AI098122 from the National Institutes of Health NIAID to KEG.

Conflict of interest

The authors claim no conflict of interest.

Supplementary material

253_2015_6443_MOESM1_ESM.pdf (931 kb)
ESM 1 (PDF 930 kb)

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Copyright information

© Springer-Verlag Berlin Heidelberg 2015

Authors and Affiliations

  • Daniel C. Osipovitch
    • 1
  • Sophie Therrien
    • 2
  • Karl E. Griswold
    • 3
    • 4
  1. 1.Program in Experimental and Molecular MedicineDartmouth CollegeHanoverUSA
  2. 2.New England CollegeHennikerUSA
  3. 3.Thayer School of EngineeringDartmouth CollegeHanoverUSA
  4. 4.Program in Molecular and Cellular BiologyDartmouth CollegeHanoverUSA

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