title: Antimicrobial protein rBPI21-induced surface changes on Gram-negative and Gram-positive bacteria.
abstract: New classes of antibiotics, such as antimicrobial peptides or proteins (AMPs), are crucial to deal with threatening bacterial diseases. rBPI21 is an AMP based on the human neutrophil BPI protein, with potential clinical use against meningitis. We studied the membrane perturbations promoted by rBPI21 on Gram-negative Escherichia coli and Gram-positive Staphylococcus aureus. Its interaction with bacteria was also studied in the presence of lipopolysaccharide (LPS), rBPI21 major ligand. Flow cytometry analysis of both bacteria shows that rBPI21 induces membrane depolarization. rBPI21 increases the negative surface charge of both bacteria toward positive values, as shown by zeta-potential measurements. This is followed by surface perturbations, culminating in cell lysis, as visualized by atomic force microscopy (AFM). Force spectroscopy measurements show that soluble LPS decreases the interaction of rBPI21 with bacteria, especially with S. aureus. This suggests that the rBPI21 LPS-binding pocket may also participate on the binding to Gram-positive bacteria.
  
  
  BACKGROUND
  In this study, rBPI21, an antimicrobial protein based on the human neutrophil BPI protein, with potential clinical use against meningitis, is analyzed with multiple tools including zeta-potential measurements, clarifying its actions on E. coli and S. aureus. Since antimicrobial peptides are potentially important new additions to antibiotic regimens, studies like this represent important cornerstones in efficiency and mechanism of action testing of these new approaches.
authors.affiliation: Instituto de Medicina Molecular, Faculdade de Medicina, Universidade de Lisboa, Lisbon, Portugal.
authors.forename: Marco M
authors.initials: MM
authors.lastname: Domingues
authors.affiliation: Instituto de Medicina Molecular, Faculdade de Medicina, Universidade de Lisboa, Lisbon, Portugal.
authors.forename: Patrícia M
authors.initials: PM
authors.lastname: Silva
authors.affiliation: Instituto de Medicina Molecular, Faculdade de Medicina, Universidade de Lisboa, Lisbon, Portugal.
authors.forename: Henri G
authors.initials: HG
authors.lastname: Franquelim
authors.affiliation: Instituto de Medicina Molecular, Faculdade de Medicina, Universidade de Lisboa, Lisbon, Portugal.
authors.forename: Filomena A
authors.initials: FA
authors.lastname: Carvalho
authors.affiliation: Instituto de Medicina Molecular, Faculdade de Medicina, Universidade de Lisboa, Lisbon, Portugal.
authors.forename: Miguel A R B
authors.initials: MA
authors.lastname: Castanho
authors.affiliation: Instituto de Medicina Molecular, Faculdade de Medicina, Universidade de Lisboa, Lisbon, Portugal. Electronic address: nsantos@fm.ul.pt.
authors.forename: Nuno C
authors.initials: NC
authors.lastname: Santos
mesh_terms: D000900:Anti-Bacterial Agents / Q000737:chemistry* / Q000494:pharmacology*; D023181:Antimicrobial Cationic Peptides / Q000737:chemistry* / Q000494:pharmacology*; D001798:Blood Proteins / Q000737:chemistry* / Q000494:pharmacology*; D004926:Escherichia coli / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D004927:Escherichia coli Infections / Q000188:drug therapy / Q000382:microbiology; D006801:Humans; D008070:Lipopolysaccharides / Q000378:metabolism; D010455:Peptides / Q000737:chemistry / Q000494:pharmacology; D011994:Recombinant Proteins / Q000737:chemistry / Q000494:pharmacology; D013203:Staphylococcal Infections / Q000188:drug therapy / Q000382:microbiology; D013211:Staphylococcus aureus / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism
chemical_list: D000900:Anti-Bacterial Agents; D023181:Antimicrobial Cationic Peptides; D001798:Blood Proteins; D008070:Lipopolysaccharides; D010455:Peptides; D011994:Recombinant Proteins; C021623:bactericidal permeability increasing protein
keywords: Lipopolysaccharide; atomic force microscopy; rBPI(21)