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Examination of yield, bacteriolytic activity and cold storage of linker deletion mutants based on endolysin S6_ORF93

Sosuke Munetomo1, Jumpei Uchiyama2, Iyo Takemura-Uchiyama2

  • 1Department of Public Health, Graduate School of Medicine Dentistry and Pharmaceutical Sciences, Okayama University, Kita-ku, Okayama, Japan.

Plos One
|October 23, 2024
PubMed

Insights

Shortening the linker in Staphylococcus endolysin ORF93 enhanced its bacteriolytic activity and protein yield. This research highlights linker optimization for developing novel antimicrobial agents against resistant bacteria.

Area of Science:

  • Microbiology
  • Biochemistry
  • Antimicrobial Research

Background:

  • Methicillin-resistant Staphylococcus species pose significant clinical and veterinary challenges due to limited effective treatments.
  • Phage-derived endolysins are promising antimicrobial agents, with their activity potentially modulated by linker regions.
  • Staphylococcus phage S6 endolysin S6_ORF93 (ORF93) was hypothesized to possess a catalytic domain, a peptidoglycan-binding domain, and a long linker.

Purpose of the Study:

  • To investigate the impact of linker length reduction on the biochemical properties and lytic activity of the Staphylococcus endolysin ORF93.
  • To engineer and characterize ORF93 linker deletion mutants for potential therapeutic applications.

Main Methods:

  • Wild-type ORF93 and four linker deletion mutants (ORF93-Δ05, ORF93-Δ10, ORF93-Δ15, ORF93-Δ20) were produced in an Escherichia coli expression system.
  • Solubility, protein yield, bacteriolytic activity, antimicrobial spectrum, and stability under cold storage were assessed for each variant.

Main Results:

  • Most mutants, except ORF93-Δ20, were successfully expressed as soluble proteins.
  • ORF93-Δ15 exhibited the highest protein yield and significant bacteriolytic activity against Staphylococcus strains.
  • The antimicrobial spectrum remained consistent across the tested variants, and linker deletion had a minimal effect on cold storage stability.

Conclusions:

  • Linker length is a critical factor in optimizing the efficacy of endolysin ORF93 as an antimicrobial agent.
  • The ORF93-Δ15 mutant demonstrates potential as a novel therapeutic candidate for combating Staphylococcus infections.
  • Further investigation into endolysin linker engineering is essential for developing effective phage-based antimicrobials.

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