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Phage-encoded cationic antimicrobial peptide required for lysis.

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|August 2, 2021
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Bacteriophage phiKT uses a novel antimicrobial peptide, a disruptin, to lyse bacterial outer membranes, offering a new lysis mechanism. This genetically tractable protein aids in studying antimicrobial peptide membrane disruption.

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Area of Science:

  • Microbiology
  • Molecular Biology
  • Biochemistry

Background:

  • Bacterial outer membrane (OM) lysis by phages typically requires spanins.
  • Approximately 15% of Gram-negative phages lack spanin genes, suggesting alternative OM disruption mechanisms.
  • The T7-like coliphage phiKT lacks spanins but exhibits explosive cell lysis.

Purpose of the Study:

  • To investigate the mechanism of outer membrane disruption in phiKT, a spanin-deficient phage.
  • To identify and characterize the novel lysis protein responsible for OM disruption.
  • To establish a new class of phage lysis proteins.

Main Methods:

  • Cloning and induction of the phiKT lysis cassette in E. coli K-12.
  • Complementation assays using a spanin-null lambda lysogen.
  • Biochemical characterization of the putative lysis protein (gp28), including urea/KCl washes.
  • Fluorescence microscopy to determine protein localization.
  • Comparison of gp28 with human cathelicidin antimicrobial peptide LL-37.

Main Results:

  • A novel gene, gp28, located between holin and endolysin genes, was identified and essential for lysis.
  • Absence of gp28 resulted in stabilized spherical cells, not lysis, indicating its role in OM disruption.
  • Gp28, a 56-amino acid cationic protein, associates with the membrane post-lysis and exhibits strong hydrophobic interactions.
  • Gp28 demonstrated functional similarities to antimicrobial peptides like LL-37.
  • Gp28 was shown to complement spanin-null phage lysis defects.

Conclusions:

  • PhiKT utilizes a phage-encoded cationic antimicrobial peptide, termed a disruptin, for outer membrane disruption during host lysis.
  • Disruptins represent a new class of phage lysis proteins with no known homologs.
  • The phiKT disruptin provides a genetically tractable system for studying antimicrobial peptide-mediated membrane disruption mechanisms.