{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,5,13]],"date-time":"2026-05-13T22:12:56Z","timestamp":1778710376604,"version":"3.51.4"},"reference-count":68,"publisher":"MDPI AG","issue":"17","license":[{"start":{"date-parts":[[2021,9,2]],"date-time":"2021-09-02T00:00:00Z","timestamp":1630540800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"Horizon 2020","award":["grant no. 685778"],"award-info":[{"award-number":["grant no. 685778"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["IJMS"],"abstract":"<jats:p>Clostridium botulinum is a Gram-positive, anaerobic, spore-forming bacterium capable of producing botulinum toxin and responsible for botulism of humans and animals. Phage-encoded enzymes called endolysins, which can lyse bacteria when exposed externally, have potential as agents to combat bacteria of the genus Clostridium. Bioinformatics analysis revealed in the genomes of several Clostridium species genes encoding putative N-acetylmuramoyl-l-alanine amidases with anti-clostridial potential. One such enzyme, designated as LysB (224-aa), from the prophage of C. botulinum E3 strain Alaska E43 was chosen for further analysis. The recombinant 27,726 Da protein was expressed and purified from E. coli Tuner(DE3) with a yield of 37.5 mg per 1 L of cell culture. Size-exclusion chromatography and analytical ultracentrifugation experiments showed that the protein is dimeric in solution. Bioinformatics analysis and results of site-directed mutagenesis studies imply that five residues, namely H25, Y54, H126, S132, and C134, form the catalytic center of the enzyme. Twelve other residues, namely M13, H43, N47, G48, W49, A50, L73, A75, H76, Q78, N81, and Y182, were predicted to be involved in anchoring the protein to the lipoteichoic acid, a significant component of the Gram-positive bacterial cell wall. The LysB enzyme demonstrated lytic activity against bacteria belonging to the genera Clostridium, Bacillus, Staphylococcus, and Deinococcus, but did not lyse Gram-negative bacteria. Optimal lytic activity of LysB occurred between pH 4.0 and 7.5 in the absence of NaCl. This work presents the first characterization of an endolysin derived from a C. botulinum Group II prophage, which can potentially be used to control this important pathogen.<\/jats:p>","DOI":"10.3390\/ijms22179536","type":"journal-article","created":{"date-parts":[[2021,9,2]],"date-time":"2021-09-02T10:02:11Z","timestamp":1630576931000},"page":"9536","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":9,"title":["Novel Lytic Enzyme of Prophage Origin from Clostridium botulinum E3 Strain Alaska E43 with Bactericidal Activity against Clostridial Cells"],"prefix":"10.3390","volume":"22","author":[{"given":"Agnieszka","family":"Morzywolek","sequence":"first","affiliation":[{"name":"Laboratory of Extremophiles Biology, Department of Microbiology, Faculty of Biology, University of Gdansk, 80-822 Gdansk, Poland"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2871-8060","authenticated-orcid":false,"given":"Magdalena","family":"Plotka","sequence":"additional","affiliation":[{"name":"Laboratory of Extremophiles Biology, Department of Microbiology, Faculty of Biology, University of Gdansk, 80-822 Gdansk, Poland"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-6598-8449","authenticated-orcid":false,"given":"Anna-Karina","family":"Kaczorowska","sequence":"additional","affiliation":[{"name":"Collection of Plasmids and Microorganisms, Faculty of Biology, University of Gdansk, 80-308 Gdansk, Poland"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Monika","family":"Szadkowska","sequence":"additional","affiliation":[{"name":"Laboratory of Extremophiles Biology, Department of Microbiology, Faculty of Biology, University of Gdansk, 80-822 Gdansk, Poland"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-8187-1980","authenticated-orcid":false,"given":"Lukasz P.","family":"Kozlowski","sequence":"additional","affiliation":[{"name":"Institute of Informatics, Faculty of Mathematics, Informatics and Mechanics, University of Warsaw, 02-097 Warsaw, Poland"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3823-3286","authenticated-orcid":false,"given":"Dariusz","family":"Wyrzykowski","sequence":"additional","affiliation":[{"name":"Department of General and Inorganic Chemistry, Faculty of Chemistry, University of Gdansk, 80-308 Gdansk, Poland"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-5283-159X","authenticated-orcid":false,"given":"Joanna","family":"Makowska","sequence":"additional","affiliation":[{"name":"Department of General and Inorganic Chemistry, Faculty of Chemistry, University of Gdansk, 80-308 Gdansk, Poland"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Jerel J.","family":"Waters","sequence":"additional","affiliation":[{"name":"Animal Biosciences and Biotechnology Laboratory, ARS, NEA, USDA, Beltsville, MD 20705-2350, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Steven M.","family":"Swift","sequence":"additional","affiliation":[{"name":"Animal Biosciences and Biotechnology Laboratory, ARS, NEA, USDA, Beltsville, MD 20705-2350, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"David M.","family":"Donovan","sequence":"additional","affiliation":[{"name":"Animal Biosciences and Biotechnology Laboratory, ARS, NEA, USDA, Beltsville, MD 20705-2350, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Tadeusz","family":"Kaczorowski","sequence":"additional","affiliation":[{"name":"Laboratory of Extremophiles Biology, Department of Microbiology, Faculty of Biology, University of Gdansk, 80-822 Gdansk, Poland"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2021,9,2]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"11","DOI":"10.1080\/10408410590912952","article-title":"Clostridium botulinum: A bug with beauty and weapon","volume":"31","author":"Shukla","year":"2005","journal-title":"Crit. 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