caa a22 4500 605506558 CHVBK 20210128100627.0 cr unu---uuuuu 210128e20150801xx s 000 0 eng 10.1007/s00253-015-6443-2 doi (NATIONALLICENCE)springer-10.1007/s00253-015-6443-2 Discovery of novel S. aureus autolysins and molecular engineering to enhance bacteriolytic activity [Elektronische Daten] [Daniel Osipovitch, Sophie Therrien, Karl Griswold] 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. Springer-Verlag Berlin Heidelberg, 2015 MRSA nationallicence Lysins nationallicence Lytic enzyme nationallicence CHAP nationallicence Lysostaphin nationallicence Antibiotic nationallicence Osipovitch Daniel Program in Experimental and Molecular Medicine, Dartmouth College, 03755, Hanover, NH, USA aut Therrien Sophie New England College, 03242, Henniker, NH, USA aut Griswold Karl Thayer School of Engineering, Dartmouth College, 03755, Hanover, NH, USA aut Applied Microbiology and Biotechnology Springer Berlin Heidelberg 99/15(2015-08-01), 6315-6326 0175-7598 99:15<6315 2015 99 253 https://doi.org/10.1007/s00253-015-6443-2 text/html Onlinezugriff via DOI BK010053 XK010053 XK010000 Metadata rights reserved Springer special CC-BY-NC licence nationallicence 1 research-article jats NATIONALLICENCE NATIONALLICENCE NL-springer NATIONALLICENCE 856 40 https://doi.org/10.1007/s00253-015-6443-2 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Osipovitch Daniel Program in Experimental and Molecular Medicine, Dartmouth College, 03755, Hanover, NH, USA aut NATIONALLICENCE 700 1- Therrien Sophie New England College, 03242, Henniker, NH, USA aut NATIONALLICENCE 700 1- Griswold Karl Thayer School of Engineering, Dartmouth College, 03755, Hanover, NH, USA aut NATIONALLICENCE 773 0- Applied Microbiology and Biotechnology Springer Berlin Heidelberg 99/15(2015-08-01), 6315-6326 0175-7598 99:15<6315 2015 99 253