title: Protecting Gram-negative bacterial cell envelopes from human lysozyme: Interactions with Ivy inhibitor proteins from Escherichia coli and Pseudomonas aeruginosa.
abstract: Lysozymes play an important role in host defense by degrading peptidoglycan in the cell envelopes of pathogenic bacteria. Several Gram-negative bacteria can evade this mechanism by producing periplasmic proteins that inhibit the enzymatic activity of lysozyme. The Escherichia coli inhibitor of vertebrate lysozyme, Ivyc and its Pseudomonas aeruginosa homolog, Ivyp1 have been shown to be potent inhibitors of hen egg white lysozyme (HEWL). Since human lysozyme (HL) plays an important role in the innate immune response, we have examined the binding of HL to Ivyc and Ivyp1. Our results show that Ivyp1 is a weaker inhibitor of HL than Ivyc even though they inhibit HEWL with similar potency. Calorimetry experiments confirm that Ivyp1 interacts more weakly with HL than HEWL. Analytical ultracentrifugation studies revealed that Ivyp1 in solution is a monomer and forms a 30kDa heterodimer with both HL and HEWL, while Ivyc is a homodimer that forms a tetramer with both enzymes. The interaction of Ivyp1 with HL was further characterized by NMR chemical shift perturbation experiments. In addition to the characteristic His-containing Ivy inhibitory loop that binds into the active site of lysozyme, an extended loop (P2) between the final two beta-strands also participates in forming protein-protein interactions. The P2 loop is not conserved in Ivyc and it constitutes a flexible region in Ivyp1 that becomes more rigid in the complex with HL. We conclude that differences in the electrostatic interactions at the binding interface between Ivy inhibitors and distinct lysozymes determine the strength of this interaction. This article is part of a Special Issue entitled: Bacterial Resistance to Antimicrobial Peptides.
authors.affiliation: Biochemistry Research Group, Department of Biological Sciences, University of Calgary, Calgary, Alberta T2N 1N4, Canada.
authors.forename: Zhihong
authors.initials: Z
authors.lastname: Liu
authors.affiliation: Biochemistry Research Group, Department of Biological Sciences, University of Calgary, Calgary, Alberta T2N 1N4, Canada.
authors.forename: Beatriz
authors.initials: B
authors.lastname: García-Díaz
authors.affiliation: Dipartimento di Scienze Biochimiche, Istituto di Biologia e Patologia Molecolari CNR, Università Sapienza, P.le A. Moro, 5-00185 Roma, Italy.
authors.forename: Bruno
authors.initials: B
authors.lastname: Catacchio
authors.affiliation: Dipartimento di Scienze Biochimiche, Istituto di Biologia e Patologia Molecolari CNR, Università Sapienza, P.le A. Moro, 5-00185 Roma, Italy.
authors.forename: Emilia
authors.initials: E
authors.lastname: Chiancone
authors.affiliation: Biochemistry Research Group, Department of Biological Sciences, University of Calgary, Calgary, Alberta T2N 1N4, Canada. Electronic address: vogel@ucalgary.ca.
authors.forename: Hans J
authors.initials: HJ
authors.lastname: Vogel
mesh_terms: D001665:Binding Sites; D002152:Calorimetry, Differential Scanning; D002352:Carrier Proteins / Q000737:chemistry / Q000276:immunology / Q000378:metabolism*; D004926:Escherichia coli / Q000276:immunology / Q000378:metabolism* / Q000472:pathogenicity; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000276:immunology / Q000378:metabolism*; D054884:Host-Pathogen Interactions; D006801:Humans; D007113:Immunity, Innate; D008958:Models, Molecular; D009113:Muramidase / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000276:immunology / Q000378:metabolism*; D019906:Nuclear Magnetic Resonance, Biomolecular; D011485:Protein Binding; D011487:Protein Conformation; D011550:Pseudomonas aeruginosa / Q000276:immunology / Q000378:metabolism* / Q000472:pathogenicity; D055672:Static Electricity; D014461:Ultracentrifugation
chemical_list: D002352:Carrier Proteins; D029968:Escherichia coli Proteins; C429555:Ivy protein, E coli; C421864:hen egg lysozyme; D009113:Muramidase
keywords: Bacterial survival; Inhibitor of vertebrate lysozyme; Ivy; Lysozyme inhibition