Production and characterization of a phage endolysin with putative antimicrobial activity against Cutibacterium acnes

<p>Acne vulgaris (acne) is the eighth most common skin disorder worldwide. Because of the heterogeneous pathology of acne and the increasing antibiotic resistance of the causal agent (Cutibacterium acnes), novel therapeutics need to be developed to treat acne. One option is endolysins, highly conserved enzymes from bacteriophages that disrupt the bacterial cell wall. Endolysins retain bactericidal and bacteriostatic activity when applied to bacteria in vitro and are safe for topical application. We hypothesize that the endolysin (ENDL) from the P100.1 C. acnes bacteriophage will exhibit bacteriostatic activity against C. acnes in vitro. This study aims to bioinformatically characterize the P100.1 endolysin’s structure, produce and isolate the endolysin, then evaluate putative antimicrobial effects. We cloned the endolysin gene into vectors (pET11a & pBAD-GFP) for expression by Escherichia coli or as a fusion protein to green fluorescent protein (GFP). We compared the growth rates of C. acnes treated with cell lysate of cultures induced to produce the endolysin to non-treatment controls to evaluate bactericidal effects. We found >88% sequence similarity between endolysins of phages targeting C. acnes, indicating high conservation and that all currently identified endolysins of phages targeting C. acnes are amidases. Expression of the recombinant proteins could only be detected for the strains containing the pBAD-GFP vector and not the pET11a vector. The antimicrobial activity of endolysin-fusion protein was inconclusive, as the impact on the growth of C. acnes over time was negligible. Therefore, more research is needed to determine whether the P100.1 endolysin or other endolysins from C. acnes phages are viable anti-bacterial therapies.</p>

Name thesis:degree_name

Master of Science (MS) in Biology

Level thesis:degree_level

Thesis

Discipline thesis:degree_discipline

Biology

Year

2023

Author dc:creator

• Collins, Kathryn M.

• Enzymes and Coenzymes
• Skin and Connective Tissue Diseases

Statement dc:rights

• Access is available to all users

Repository record dc:identifier

https://dc.ewu.edu/theses/839

OAI identifier oai:identifier

oai:dc.ewu.edu:theses-1841