{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,25]],"date-time":"2026-07-25T02:19:19Z","timestamp":1784945959145,"version":"3.55.0"},"reference-count":35,"publisher":"MDPI AG","issue":"3","license":[{"start":{"date-parts":[[2023,3,4]],"date-time":"2023-03-04T00:00:00Z","timestamp":1677888000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100003725","name":"National Research Foundation of Korea","doi-asserted-by":"publisher","award":["NRF-2019M3E5D5066666"],"award-info":[{"award-number":["NRF-2019M3E5D5066666"]}],"id":[{"id":"10.13039\/501100003725","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100003725","name":"National Research Foundation of Korea","doi-asserted-by":"publisher","award":["HI21C2447"],"award-info":[{"award-number":["HI21C2447"]}],"id":[{"id":"10.13039\/501100003725","id-type":"DOI","asserted-by":"publisher"}]},{"name":"Korea Health Industry Development Institute (KHIDI)","award":["NRF-2019M3E5D5066666"],"award-info":[{"award-number":["NRF-2019M3E5D5066666"]}]},{"name":"Korea Health Industry Development Institute (KHIDI)","award":["HI21C2447"],"award-info":[{"award-number":["HI21C2447"]}]},{"name":"HUFS Research Fund of 2023","award":["NRF-2019M3E5D5066666"],"award-info":[{"award-number":["NRF-2019M3E5D5066666"]}]},{"name":"HUFS Research Fund of 2023","award":["HI21C2447"],"award-info":[{"award-number":["HI21C2447"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Viruses"],"abstract":"<jats:p>Genes encoding endolysins were identified and cloned from three different Escherichia coli bacteriophages, 10-24(13), PBEC30, and PBEC56. Putative antimicrobial peptide (AMP)-like C-terminal alpha helix structures with amphipathic natures were predicted from the three endolysins. Each gene was cloned and expressed as hexahistidine-tagged forms, and the products were purified and characterized. The purified endolysins exhibited antibacterial activities against a variety of Gram-negative bacteria including Escherichia coli, Pseudomonas aeruginosa, Acinetobacter baumannii, and Klebsiella pneumonia. Their antibacterial activities were improved by N-terminal fusion with an antimicrobial peptide, cecropin A. Minimum inhibitory concentrations (MIC) were as low as 4 \u03bcg\/mL, depending on the targeted strain. The endolysins\u2019 enzymatic activities were not affected by changes in pH at ranges from 5 to 10 and were stable at temperatures between 4 and 65 \u00b0C. The in vivo efficacies of the three endolysins were also demonstrated using Galleria melonella for infection models.<\/jats:p>","DOI":"10.3390\/v15030679","type":"journal-article","created":{"date-parts":[[2023,3,6]],"date-time":"2023-03-06T05:18:43Z","timestamp":1678079923000},"page":"679","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":25,"title":["Characterization of Three Different Endolysins Effective against Gram-Negative Bacteria"],"prefix":"10.3390","volume":"15","author":[{"given":"Tae-Hwan","family":"Jeong","sequence":"first","affiliation":[{"name":"Department of Bioscience and Biotechnology, Hankuk University of Foreign Studies, Yong-In 17035, Gyung-Gi Do, Republic of Korea"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Hye-Won","family":"Hong","sequence":"additional","affiliation":[{"name":"LyseNTech Co. Ltd., Seongnam 13486, Gyung-Gi Do, Republic of Korea"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Min Soo","family":"Kim","sequence":"additional","affiliation":[{"name":"LyseNTech Co. Ltd., Seongnam 13486, Gyung-Gi Do, Republic of Korea"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Miryoung","family":"Song","sequence":"additional","affiliation":[{"name":"Department of Bioscience and Biotechnology, Hankuk University of Foreign Studies, Yong-In 17035, Gyung-Gi Do, Republic of Korea"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7478-7676","authenticated-orcid":false,"given":"Heejoon","family":"Myung","sequence":"additional","affiliation":[{"name":"Department of Bioscience and Biotechnology, Hankuk University of Foreign Studies, Yong-In 17035, Gyung-Gi Do, Republic of Korea"},{"name":"LyseNTech Co. Ltd., Seongnam 13486, Gyung-Gi Do, Republic of Korea"},{"name":"The Bacteriophage Bank of Korea, Seongnam 13486, Gyung-Gi Do, Republic of Korea"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2023,3,4]]},"reference":[{"key":"ref_1","unstructured":"Antimicrobial Resistance Collaborators (2022). Global Burden of Bacterial Antimicrobial Resistance in 2019: A Systematic Analysis. Lancet, 399, 629."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"822689","DOI":"10.3389\/fmicb.2022.822689","article-title":"A Review of Current Bacterial Resistance to Antibiotics in Food Animals","volume":"13","author":"Xu","year":"2022","journal-title":"Front. Microbiol."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"735","DOI":"10.2147\/IDR.S345574","article-title":"Clinical Perspective of Antimicrobial Resistance in Bacteria","volume":"15","author":"Zhu","year":"2022","journal-title":"Infect. Drug Resist."},{"key":"ref_4","first-page":"e00181","article-title":"Antimicrobial Resistance in ESKAPE Pathogens","volume":"33","author":"Forde","year":"2020","journal-title":"Clin. Microbiol. Rev."},{"key":"ref_5","first-page":"726","article-title":"Towards the sustainable discovery and development of new antibiotics","volume":"5","author":"Miethke","year":"2021","journal-title":"Nature"},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"415","DOI":"10.2147\/IDR.S287792","article-title":"Encouraging the Development of New Antibiotics: Are Financial Incentives the Right Way Forward? A Systematic Review and Case Study","volume":"14","author":"Dutescu","year":"2021","journal-title":"Infect. Drug Resist."},{"key":"ref_7","first-page":"e00071","article-title":"Recombinant Endolysins as Potential Therapeutics against Antibiotic-Resistant Staphylococcus aureus: Current Status of Research and Novel Delivery Strategies","volume":"31","author":"Kashani","year":"2017","journal-title":"Clin. Microbiol. Rev."},{"key":"ref_8","doi-asserted-by":"crossref","unstructured":"Murray, E., Draper, L.A., Ross, R.P., and Hill, C. (2021). The Advantages and Challenges of using Endolysins in a Clinical Setting. Viruses, 13.","DOI":"10.3390\/v13040680"},{"key":"ref_9","doi-asserted-by":"crossref","unstructured":"Rahman, M.u., Wang, W., Sun, Q., Shah, J.A., Li, C., Sun, Y., Li, Y., Zhang, B., Chen, W., and Wang, S. (2021). Endolysin, a Promising Solution against Antimicrobial Resistance. Antibiotics, 10.","DOI":"10.3390\/antibiotics10111277"},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"243","DOI":"10.1007\/s12275-014-4087-z","article-title":"Phage lysis: Three steps, three choices, one outcome","volume":"52","author":"Young","year":"2014","journal-title":"J. Microbiol."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"2081","DOI":"10.1021\/acsinfecdis.0c00812","article-title":"Development of Advanced Chimeric Endolysin to Control Multidrug-Resistant Staphylococcus aureus through Domain Shuffling","volume":"7","author":"Lee","year":"2021","journal-title":"ACS Infect. Dis."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"668430","DOI":"10.3389\/fcimb.2021.668430","article-title":"Phage Endolysin LysP108 showed Promising Antibacterial Potential against Methicillin-Resistant Staphylococcus aureus","volume":"11","author":"Lu","year":"2021","journal-title":"Front. Cell Infect. Microbiol."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"1916","DOI":"10.4014\/jmb.1908.08021","article-title":"Characteristics and Lytic Activity of Phage-Derived Peptidoglycan Hydrolase, LysSAP8, as a Potent Alternative Biocontrol Agent for Staphylococcus aureus","volume":"29","author":"Yu","year":"2019","journal-title":"J. Microbiol. Biotechnol."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"e00731","DOI":"10.1128\/AAC.00731-18","article-title":"Effects of Phage Endolysin SAL200 Combined with Antibiotics on Staphylococcus aureus Infection","volume":"62","author":"Kim","year":"2018","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_15","doi-asserted-by":"crossref","unstructured":"Zhang, W., Mi, Z., Yin, X., Fan, H., An, X., Zhang, Z., Chen, J., and Tong, Y. (2013). Characterization of Enterococcus faecalis Phage IME-EF1 and its Endolysin. PLoS ONE, 8.","DOI":"10.1371\/journal.pone.0080435"},{"key":"ref_16","doi-asserted-by":"crossref","unstructured":"Son, B., Yun, J., Lim, J., Shin, H., Heu, S., and Ryu, S. (2012). Characterization of LysB4, an Endolysin from the Bacillus cereus-Infecting Bacteriophage B4. BMC Microbiol., 12.","DOI":"10.1186\/1471-2180-12-33"},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"293","DOI":"10.3389\/fmicb.2017.00293","article-title":"A Novel Antimicrobial Endolysin, LysPA26, against Pseudomonas aeruginosa","volume":"8","author":"Guo","year":"2017","journal-title":"Front. Microbiol."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"3302","DOI":"10.3389\/fmicb.2018.03302","article-title":"A Novel Phage PD-6A3, and its Endolysin Ply6A3, with Extended Lytic Activity against Acinetobacter baumannii","volume":"9","author":"Wu","year":"2018","journal-title":"Front. Microbiol."},{"key":"ref_19","doi-asserted-by":"crossref","unstructured":"Plotka, M., Kapusta, M., Dorawa, S., Kaczorowska, A., and Kaczorowski, T. (2019). Ts2631 Endolysin from the Extremophilic Thermus Scotoductus Bacteriophage vB_Tsc2631 as an Antimicrobial Agent against Gram-Negative Multidrug-Resistant Bacteria. Viruses, 11.","DOI":"10.3390\/v11070657"},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"32","DOI":"10.1016\/j.jgar.2020.01.005","article-title":"Antimicrobial Activity of LysSS, a Novel Phage Endolysin, against Acinetobacter baumannii and Pseudomonas aeruginosa","volume":"22","author":"Kim","year":"2020","journal-title":"J. Glob. Antimicrob. Rsist."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"637313","DOI":"10.3389\/fcimb.2021.637313","article-title":"A Novel Acinetobacter baumannii Bacteriophage Endolysin LysAB54 with High Antibacterial Activity against Multiple Gram-Negative Microbes","volume":"11","author":"Khan","year":"2021","journal-title":"Front. Cell. Infect. Microbiol."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"3774","DOI":"10.1128\/AAC.02668-14","article-title":"Art-175 is a Highly Efficient Antibacterial against Multidrug-Resistant Strains and Persisters of Pseudomonas aeruginosa","volume":"58","author":"Briers","year":"2014","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"3480","DOI":"10.1128\/AAC.00285-16","article-title":"Efficacy of Artilysin Art-175 against Resistant and Persistent Acinetobacter baumannii","volume":"60","author":"Defraine","year":"2016","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"e0151521","DOI":"10.1128\/AEM.01515-21","article-title":"Engineering a Lysin with Intrinsic Antibacterial Activity (LysMK34) by Cecropin A Fusion Enhances its Antibacterial Properties against Acinetobacter baumannii","volume":"88","author":"Abdelkader","year":"2022","journal-title":"Appl. Environ. Microbiol."},{"key":"ref_25","doi-asserted-by":"crossref","unstructured":"Antonova, N.P., Vasina, D.V., Lendel, A.M., Usachev, E.V., Makarov, V.V., Gintsburg, A.L., Tkachuk, A.P., and Gushchin, V.A. (2019). Broad Bactericidal Activity of the Myoviridae Bacteriophage Lysins LysAm24, LysECD7, and LysSi3 against Gram-Negative ESKAPE Pathogens. Viruses, 11.","DOI":"10.3390\/v11030284"},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"816","DOI":"10.4014\/jmb.2205.05009","article-title":"Bactericidal Effect of Cecropin A Fused Endolysin on Drug-Resistant Gram-Negative Pathogens","volume":"32","author":"Lim","year":"2022","journal-title":"J. Microbiol. Biotechnol."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"859","DOI":"10.1007\/s12275-022-2107-y","article-title":"Eradication of drug-resistant Acinetobacter baumannii by cell-penetrating peptide fused endolysin","volume":"60","author":"Lim","year":"2022","journal-title":"J. Microbiol."},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"821936","DOI":"10.3389\/fmicb.2022.821936","article-title":"Combination Effect of Engineered Endolysin EC340 with Antibiotics","volume":"13","author":"Hong","year":"2022","journal-title":"Front. Microbiol."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"e0032121","DOI":"10.1128\/JVI.00321-21","article-title":"Sequence-Function Relationships in Phage-Encoded Bacterial Cell Wall Lytic Enzymes and Their Implications for Phage-Derived Product Design","volume":"95","year":"2021","journal-title":"J. Virol."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"15","DOI":"10.1016\/j.copbio.2020.08.014","article-title":"Lysins Breaking Down the Walls of Gram-Negative Bacteria, no Longer a no-Go","volume":"68","author":"Briers","year":"2021","journal-title":"Curr. Opin. Biotechnol."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"e00342","DOI":"10.1128\/AAC.00342-19","article-title":"Lysocins: Bioengineered Antimicrobials that Deliver Lysins Across the Outer Membrane of Gram-Negative Bacteria","volume":"63","author":"Heselpoth","year":"2019","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"602","DOI":"10.1128\/AAC.44.3.602-607.2000","article-title":"Antibacterial and Antimembrane Activities of Cecropin A in Escherichia coli","volume":"44","author":"Silvestro","year":"2000","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"3864","DOI":"10.1039\/c2ra20140b","article-title":"Tuning the Bactericidal Repertoire and Potency of Quinoline-Based Amphiphiles for Enhanced Killing of Pathogenic Bacteria","volume":"2","author":"Vudumula","year":"2012","journal-title":"RSC Adv."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"1750","DOI":"10.1016\/j.bpj.2016.09.001","article-title":"Bilayer Properties of Lipid A from various Gram-Negative Bacteria","volume":"111","author":"Kim","year":"2016","journal-title":"Biophys. J."},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"16062","DOI":"10.1021\/acsami.1c01643","article-title":"Structural Disruptions of the Outer Membranes of Gram-Negative Bacteria by Rationally Designed Amphiphilic Antimicrobial Peptides","volume":"13","author":"Gong","year":"2021","journal-title":"ACS Appl. Mater. Interfaces"}],"container-title":["Viruses"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1999-4915\/15\/3\/679\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,10]],"date-time":"2025-10-10T18:47:46Z","timestamp":1760122066000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1999-4915\/15\/3\/679"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2023,3,4]]},"references-count":35,"journal-issue":{"issue":"3","published-online":{"date-parts":[[2023,3]]}},"alternative-id":["v15030679"],"URL":"https:\/\/doi.org\/10.3390\/v15030679","relation":{},"ISSN":["1999-4915"],"issn-type":[{"value":"1999-4915","type":"electronic"}],"subject":[],"published":{"date-parts":[[2023,3,4]]}}}