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Publication Abstract from PubMed

OBJECTIVE: Clostridioides difficile is a Gram-positive, spore-forming obligate anaerobe that can cause symptoms such as diarrhoea and abdominal cramping and lead to conditions including pseudomembranous colitis. The mainstay treatment of C. difficile infection (CDI) is antibiotics; however, antibiotics-induced gut microbiota dysbiosis poses a potential risk for refractory CDI and increased CDI recurrences. Recently, there has been growing interest in bacteriophages and their lytic enzymes as alternatives to antibiotics. We aimed to develop an engineered endolysin to specifically inactivate C. difficile. METHODS: Endolysins are bacteriolytic enzymes that facilitate the release of phage progeny during the final stage of infection. We previously demonstrated that CD27L_EAD, an engineered endolysin, inactivated C. difficile in a co-culture model of human gut microbiota. This study modified the PhiCD111 endolysin to PHICD111_20024_EAD by cleaving the cell wall-binding domain and characterized its catalytic activity in comparison with CD27L_EAD. RESULTS: PHICD111_20024_EAD shows high sequence similarity with CD27L_EAD. However, PHICD111_20024_EAD exerted superior bacteriolytic activity compared to CD27L_EAD under high salt concentrations. PHICD111_20024_EAD and CD27L_EAD displayed similar overall structures, consisting of five alpha-helices and six beta-strands, and were superimposed with a zinc ion at the active site, suggesting that both enzymes are zinc-dependent N-acetylmuramoyl-l-alanine amidases. However, their binding affinities for zinc ions differed, and excess zinc ions inhibited the catalytic activity of PHICD111_20024_EAD. CONCLUSIONS: For feasible applications of endolysins in dynamic gut environments, PHICD111_20024_EAD, which is tolerant to high osmolarity and can be modulated using zinc concentrations, provides advantageous attributes for developing therapeutic agents against CDI.

In vitro characterization of PHICD111_20024_EAD as an engineered endolysin against Clostridioides difficile.,Park J, Yoo S, An J, Cho Y, Park K, Hwang J, Chun SI, Kang NJ, Seo MD, Yoon H J Glob Antimicrob Resist. 2025 Aug 20;45:1-10. doi: 10.1016/j.jgar.2025.08.009. PMID:40845944^[1]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.