title: Enhanced antimicrobial activity of engineered human lysozyme.
abstract: Lysozymes contain a disproportionately large fraction of cationic residues, and are thereby attracted toward the negatively charged surface of bacterial targets. Importantly, this conserved biophysical property may inhibit lysozyme antibacterial function during acute and chronic infections. A mouse model of acute pulmonary Pseudomonas aeruginosa infection demonstrated that anionic biopolymers accumulate to high concentrations in the infected lung, and the presence of these species correlates with decreased endogenous lysozyme activity. To develop antibacterial enzymes designed specifically to be used as antimicrobial agents in the infected airway, the electrostatic potential of human lysozyme (hLYS) was remodeled by protein engineering. A novel, high-throughput screen was implemented to functionally interrogate combinatorial libraries of charge-engineered hLYS proteins, and variants with improved bactericidal activity were isolated and characterized in detail. These studies illustrate a general mechanism by which polyanions inhibit lysozyme function, and they are the first direct demonstration that decreasing hLYS's net cationic character improves its antibacterial activity in the presence of disease-associated biopolymers. In addition to avoiding electrostatic sequestration, at least one charge-engineered variant also kills bacteria more rapidly in the absence of inhibitory biopolymers; this observation supports a novel hypothesis that tuning the cellular affinity of peptidoglycan hydrolases may be a general strategy for improving kinetics of bacterial killing.
authors.affiliation: Dartmouth College, Hanover, New Hampshire 03755, USA.
authors.forename: Thomas C
authors.initials: TC
authors.lastname: Scanlon
authors.forename: Charlotte C
authors.initials: CC
authors.lastname: Teneback
authors.forename: Avinash
authors.initials: A
authors.lastname: Gill
authors.forename: Jenna L
authors.initials: JL
authors.lastname: Bement
authors.forename: Joshua A
authors.initials: JA
authors.lastname: Weiner
authors.forename: John W
authors.initials: JW
authors.lastname: Lamppa
authors.forename: Laurie W
authors.initials: LW
authors.lastname: Leclair
authors.forename: Karl E
authors.initials: KE
authors.lastname: Griswold
mesh_terms: D000818:Animals; D000900:Anti-Bacterial Agents / Q000737:chemistry* / Q000378:metabolism / Q000494:pharmacology / Q000627:therapeutic use*; D001419:Bacteria / Q000187:drug effects*; D006090:Gram-Negative Bacteria / Q000187:drug effects; D006801:Humans; D008168:Lung / Q000187:drug effects; D051379:Mice; D016982:Micrococcus luteus / Q000187:drug effects; D008958:Models, Molecular; D009113:Muramidase / Q000737:chemistry* / Q000235:genetics / Q000494:pharmacology / Q000627:therapeutic use*; D009154:Mutation; D015202:Protein Engineering; D011552:Pseudomonas Infections / Q000188:drug therapy*; D011550:Pseudomonas aeruginosa / Q000187:drug effects; D055672:Static Electricity
chemical_list: D000900:Anti-Bacterial Agents; D009113:Muramidase