{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,6,23]],"date-time":"2026-06-23T13:42:01Z","timestamp":1782222121370,"version":"3.54.5"},"reference-count":56,"publisher":"Frontiers Media SA","license":[{"start":{"date-parts":[[2022,7,4]],"date-time":"2022-07-04T00:00:00Z","timestamp":1656892800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100001809","name":"National Natural Science Foundation of China","doi-asserted-by":"publisher","id":[{"id":"10.13039\/501100001809","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100005046","name":"Natural Science Foundation of Heilongjiang Province","doi-asserted-by":"publisher","id":[{"id":"10.13039\/501100005046","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["frontiersin.org"],"crossmark-restriction":true},"short-container-title":["Front. Microbiol."],"abstract":"<jats:p>Antimicrobial peptides (AMPs) have attracted extensive attention because of their broad-spectrum antibacterial activity and low level of induced bacterial resistance. However, the development of some natural AMPs does not consider the perfect balance of structural characteristics, resulting in some empirical and controversial practices still existing. To further explore and complete the relationship between parameters and function of \u03b1-helix peptide, in this study, the natural antimicrobial peptide TP secreted from Bacillus strain of Tibetan pigs was selected as a template to investigate the effect of systematic mutations in the hydrogen bond formation site of the \u03b1-helical antimicrobial peptide on the activity and cell selectivity of the antimicrobial peptide. The target peptide TP(i+4) 1&amp;amp;2&amp;amp;5 with modification of two pairs of positively charged amino acids and a pair of hydrophobic amino acids showed excellent antibacterial ability and the best selectivity index (SI = 64) <jats:italic>in vitro<\/jats:italic>. At the same time, TP(i+4) 1&amp;amp;2&amp;amp;5 remained active in the presence of physiological salts and serum. The results of fluorescence, flow cytometry, and electron microscopy showed that the optimized sequences showed good antibacterial activity by membrane infiltration and membrane destruction. The potential of TP(i+4) 1&amp;amp;2&amp;amp;5 <jats:italic>in vivo<\/jats:italic> was tested in a mouse peritonitis model. Organ bacterial loads in the liver, kidney, spleen, and lungs of mice treated with TP(i+4) 1&amp;amp;2&amp;amp;5 were significantly lower compared to the infected group (<jats:italic>p<\/jats:italic> &amp;lt; 0.05). Overall, these findings contribute to the design and optimization of antimicrobial peptides with high activity and low toxicity and may accelerate the clinical application of antimicrobial peptides.<\/jats:p>","DOI":"10.3389\/fmicb.2022.864374","type":"journal-article","created":{"date-parts":[[2022,7,4]],"date-time":"2022-07-04T09:01:14Z","timestamp":1656925274000},"update-policy":"https:\/\/doi.org\/10.3389\/crossmark-policy","source":"Crossref","is-referenced-by-count":8,"title":["Optimization of Antibacterial Activity in Tibetan Swine \u03b1-Helix Peptide TP by Site-Directed Mutagenesis"],"prefix":"10.3389","volume":"13","author":[{"given":"Guoyu","family":"Li","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xiaojie","family":"Yuan","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Hongyu","family":"Chen","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Bowen","family":"Li","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Changxuan","family":"Shao","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yongjie","family":"Zhu","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Zhenheng","family":"Lai","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Anshan","family":"Shan","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1965","published-online":{"date-parts":[[2022,7,4]]},"reference":[{"key":"B1","doi-asserted-by":"publisher","first-page":"43","DOI":"10.1016\/j.drup.2016.04.002","article-title":"Mechanisms and consequences of bacterial resistance to antimicrobial peptides","volume":"26","author":"Andersson","year":"2016","journal-title":"Drug Resist. Updates"},{"key":"B2","doi-asserted-by":"publisher","first-page":"238","DOI":"10.1038\/nrmicro1098","article-title":"Antimicrobial peptides: pore formers or metabolic inhibitors in bacteria?","volume":"3","author":"Brogden","year":"2005","journal-title":"Nat. Rev. Microbiol"},{"key":"B3","doi-asserted-by":"publisher","first-page":"24","DOI":"10.1016\/j.coi.2005.11.004","article-title":"Cationic host defense (antimicrobial) peptides","volume":"18","author":"Brown","year":"2006","journal-title":"Curr. Opin. Immunol."},{"key":"B4","doi-asserted-by":"publisher","first-page":"1398","DOI":"10.1128\/AAC.00925-06","article-title":"Role of peptide hydrophobicity in the mechanism of action of alpha-helical antimicrobial peptides","volume":"51","author":"Chen","year":"2007","journal-title":"Antimicrobial Agents Chemother."},{"key":"B5","doi-asserted-by":"publisher","first-page":"4050","DOI":"10.1128\/AAC.00252-13","article-title":"Resistance of short antimicrobial peptides","volume":"57","author":"Chu","year":"2013","journal-title":"Antimicrobial Agents Chemother."},{"key":"B6","doi-asserted-by":"publisher","first-page":"2155","DOI":"10.1016\/j.bbamem.2018.08.012","article-title":"Biophysical characterization and antitumor activity of synthetic Pantinin peptides from scorpion's venom","volume":"1860","author":"Crusca","year":"2018","journal-title":"Biochim. Biophys. Acta Biomembr."},{"key":"B7","doi-asserted-by":"publisher","first-page":"254","DOI":"10.1016\/j.actbio.2021.01.036","article-title":"Binding loop of sunflower trypsin inhibitor 1 serves as a design motif for proteolysis-resistant antimicrobial peptides","volume":"124","author":"Cw","year":"2021","journal-title":"Acta Biomaterialia."},{"key":"B8","doi-asserted-by":"publisher","first-page":"3208","DOI":"10.1128\/AAC.49.8.3208-3216.2005","article-title":"Activity of the de novo engineered antimicrobial peptide WLBU2 against Pseudomonas aeruginosa in human serum and whole blood: implications for systemic applications","volume":"49","author":"Deslouches","year":"2005","journal-title":"Antimicrobial Agents Chemother."},{"key":"B9","doi-asserted-by":"publisher","first-page":"63","DOI":"10.1007\/s12154-008-0010-6","article-title":"Sterols and membrane dynamics","volume":"1","author":"Dufourc","year":"2008","journal-title":"J. Chem. Biol."},{"key":"B10","doi-asserted-by":"publisher","first-page":"3708","DOI":"10.1128\/AAC.00380-10","article-title":"Depolarization, bacterial membrane composition, and the antimicrobial action of ceragenins","volume":"54","author":"Epand","year":"2010","journal-title":"Antimicrobial Agents Chemother."},{"key":"B11","doi-asserted-by":"publisher","first-page":"e2101304","DOI":"10.1002\/smtd.202101304","article-title":"Sweeper: simplified self-assembled octapeptides with double-layered Trp zipper induces membrane destabilization and bacterial apoptosis-like death","volume":"5","author":"Fang","year":"2021","journal-title":"Small Methods"},{"key":"B12","doi-asserted-by":"publisher","first-page":"1146","DOI":"10.3389\/fmicb.2020.01146","article-title":"The critical role of tryptophan in the antimicrobial activity and cell toxicity of the duck antimicrobial peptide DCATH","volume":"11","author":"Feng","year":"2020","journal-title":"Front. Microbiol."},{"key":"B13","doi-asserted-by":"publisher","first-page":"4049","DOI":"10.1128\/AAC.00530-10","article-title":"Cationic amphiphiles, a new generation of antimicrobials inspired by the natural antimicrobial peptide scaffold","volume":"54","author":"Findlay","year":"2010","journal-title":"Antimicrobial Agents Chemother."},{"key":"B14","doi-asserted-by":"publisher","first-page":"13460","DOI":"10.1021\/acs.jafc.1c05820","article-title":"Immune defense through calcium-sensing receptor (CaSR)-dependent metabolic pathways","volume":"69","author":"Gao","year":"2021","journal-title":"J. Agric. Food Chem."},{"key":"B15","doi-asserted-by":"publisher","first-page":"34609","DOI":"10.1021\/acsami.9b10028","article-title":"Hydrophobic control of the bioactivity and cytotoxicity of de novo-designed antimicrobial peptides","volume":"11","author":"Gong","year":"2019","journal-title":"ACS Appl. Mater. Interfaces"},{"key":"B16","doi-asserted-by":"publisher","first-page":"505","DOI":"10.1007\/s00726-015-2104-0","article-title":"Design and membrane-disruption mechanism of charge-enriched AMPs exhibiting cell selectivity, high-salt resistance, anti-biofilm properties","volume":"48","author":"Han","year":"2016","journal-title":"Amino Acids"},{"key":"B17","doi-asserted-by":"publisher","first-page":"181","DOI":"10.1016\/j.intimp.2016.07.029","article-title":"Recombinant expression of porcine lactoferrin peptide LF-6 with intein technology and its immunomodulatory function in ETEC K88-infected mice","volume":"39","author":"Jiang","year":"2016","journal-title":"Int. Immunopharmacol."},{"key":"B18","doi-asserted-by":"publisher","first-page":"20150292","DOI":"10.1098\/rstb.2015.0292","article-title":"Bacterial strategies of resistance to antimicrobial peptides","volume":"371","author":"Joo","year":"2016","journal-title":"Philos. Trans. R. Soc. Lond. B Biol."},{"key":"B19","doi-asserted-by":"publisher","first-page":"2250","DOI":"10.1016\/j.bbamem.2014.05.013","article-title":"The role of spontaneous lipid curvature in the interaction of interfacially active peptides with membranes","volume":"1838","author":"Koller","year":"2014","journal-title":"Biochim. Biophys. Acta Biomembr."},{"key":"B20","doi-asserted-by":"publisher","first-page":"112580","DOI":"10.1016\/j.ejmech.2020.112580","article-title":"Antimicrobial and anti-inflammatory activities of short dodecapeptides derived from duck cathelicidin: plausible mechanism of bactericidal action and endotoxin neutralization","volume":"204","author":"Kumar","year":"2020","journal-title":"Eur. J. Med. Chem."},{"key":"B21","doi-asserted-by":"publisher","DOI":"10.1021\/acsnano.1c03301","article-title":"Self-assembling peptide Dendron nanoparticles with high stability and a multimodal antimicrobial mechanism of action","author":"Lai","year":"2021","journal-title":"ACS Nano"},{"key":"B22","doi-asserted-by":"publisher","first-page":"22113","DOI":"10.1021\/acsami.9b04654","article-title":"Highly stabilized \u03b1-helical coiled coils kill gram-negative bacteria by multicomplementary mechanisms under acidic condition","volume":"11","author":"Lai","year":"2019","journal-title":"ACS Appl. Mater. Interfaces"},{"key":"B23","doi-asserted-by":"publisher","first-page":"107962","DOI":"10.1016\/j.biotechadv.2022.107962","article-title":"Strategies employed in the design of antimicrobial peptides with enhanced proteolytic stability","volume":"59","author":"Lai","year":"2022","journal-title":"Biotechnol. Adv."},{"key":"B24","doi-asserted-by":"publisher","first-page":"e67597","DOI":"10.1371\/journal.pone.0067597","article-title":"A proline-hinge alters the characteristics of the amphipathic \u03b1-helical AMPs","volume":"8","author":"Lee","year":"2013","journal-title":"PLoS ONE"},{"key":"B25","doi-asserted-by":"publisher","first-page":"12124","DOI":"10.1038\/s41598-017-10839-4","article-title":"and immunomodulatory activities of insect defensins-DLP2 and DLP4 against multidrug-resistant Staphylococcus aureus","volume":"7","author":"Li","year":"2017","journal-title":"Sci. Rep."},{"key":"B26","doi-asserted-by":"publisher","first-page":"12313","DOI":"10.3390\/ijms140612313","article-title":"Contribution of the Tyr-1 in plantaricin149a to disrupt phospholipid model membranes","volume":"14","author":"Lopes","year":"2013","journal-title":"Int. J. Mol. Sci."},{"key":"B27","doi-asserted-by":"publisher","first-page":"374","DOI":"10.1002\/prot.23188","article-title":"Prediction of protein secondary structure from circular dichroism using theoretically derived spectra","volume":"80","author":"Louis-Jeune","year":"2012","journal-title":"Proteins Struct. Funct. Bioinformatics"},{"key":"B28","doi-asserted-by":"publisher","first-page":"e86364","DOI":"10.1371\/journal.pone.0086364","article-title":"Antimicrobial properties and membrane-active mechanism of a potential \u03b1-helical antimicrobial derived from cathelicidin PMAP-36","volume":"9","author":"Lv","year":"2014","journal-title":"PLoS ONE"},{"key":"B29","doi-asserted-by":"publisher","first-page":"6941","DOI":"10.1021\/acs.jmedchem.9b00288","article-title":"Design of Trp-rich dodecapeptides with broad-spectrum antimicrobial potency and membrane-disruptive mechanism","volume":"62","author":"Lyu","year":"2019","journal-title":"J. Med. Chem."},{"key":"B30","doi-asserted-by":"publisher","first-page":"517","DOI":"10.1016\/j.biomaterials.2015.02.063","article-title":"Characterization of cell selectivity, physiological stability and endotoxin neutralization capabilities of \u03b1-helix-based peptide amphiphiles","volume":"52","author":"Ma","year":"2015","journal-title":"Biomaterials"},{"key":"B31","doi-asserted-by":"publisher","first-page":"1193","DOI":"10.1001\/jama.2016.11764","article-title":"Antimicrobial resistance","volume":"316","author":"Marston","year":"2016","journal-title":"JAMA"},{"key":"B32","doi-asserted-by":"publisher","first-page":"201821410","DOI":"10.1073\/pnas.1821410116","article-title":"Low cationicity is important for systemic in vivo efficacy of database-derived peptides against drug-resistant Gram-positive pathogens","volume":"116","author":"Mishra","year":"2019","journal-title":"Proc. Nat. Acad. Sci. U. S. A."},{"key":"B33","doi-asserted-by":"publisher","first-page":"311","DOI":"10.1038\/s41573-019-0058-8","article-title":"Antimicrobial host defence peptides: functions and clinical potential","volume":"19","author":"Mookherjee","year":"2020","journal-title":"Nat. Rev. Drug Discov"},{"key":"B34","doi-asserted-by":"publisher","first-page":"1315","DOI":"10.1016\/j.biomaterials.2013.10.053","article-title":"of stereochemistry, chain length and sequence pattern on antimicrobial properties of short synthetic \u03b2-sheet forming peptide amphiphiles","volume":"35","author":"Ong","year":"2014","journal-title":"Biomaterials"},{"key":"B35","doi-asserted-by":"publisher","first-page":"201704489","DOI":"10.1073\/pnas.1704489114","article-title":"Lipid topology and electrostatic interactions underpin lytic activity of linear cationic antimicrobial peptides in membranes","volume":"114","author":"Paterson","year":"2017","journal-title":"Proc. Natl. Acad. Sci. U. S. A."},{"key":"B36","doi-asserted-by":"publisher","first-page":"256","DOI":"10.1016\/j.bbamem.2018.06.016","article-title":"Antimicrobial and anti-inflammatory activities of chemokine CXCL14-derived antimicrobial peptide and its analogs, Biochimica et biophysica acta","volume":"1861","author":"Rajasekaran","year":"2019","journal-title":"Biomembranes"},{"key":"B37","doi-asserted-by":"publisher","first-page":"2924","DOI":"10.1021\/acs.jmedchem.7b01831","article-title":"Antibacterial activity affected by the conformational flexibility in glycine-lysine based \u03b1-helical antimicrobial peptides","volume":"61","author":"Ron\u010devi\u0107","year":"2018","journal-title":"J. Med. Chem."},{"key":"B38","doi-asserted-by":"publisher","first-page":"e45186","DOI":"10.1371\/journal.pone.0045186","article-title":"Distinct licensing of IL-18 and IL-1\u03b2 secretion in response to NLRP3 inflammasome activation","volume":"7","author":"Schmidt","year":"2012","journal-title":"PLoS ONE"},{"key":"B39","doi-asserted-by":"publisher","first-page":"2003899","DOI":"10.1002\/smll.202003899","article-title":"Cross-strand interaction, central bending, and sequence pattern act as biomodulators of simplified \u03b2-hairpin antimicrobial amphiphiles","volume":"17","author":"Shao","year":"2020","journal-title":"Small"},{"key":"B40","doi-asserted-by":"publisher","first-page":"2440","DOI":"10.1016\/j.biomaterials.0.2005.11.036","article-title":"Antibacterial and mechanical properties of bone cement impregnated with chitosan nanoparticles","volume":"27","author":"Shi","year":"2006","journal-title":"Biomaterials"},{"key":"B41","doi-asserted-by":"publisher","first-page":"3334","DOI":"10.1021\/jm0703542","article-title":"Albumin binding of short cationic antimicrobial micropeptides and its influence on the in vitro bactericidal effect","volume":"50","author":"Svenson","year":"2007","journal-title":"J. Med. Chem."},{"key":"B42","doi-asserted-by":"publisher","first-page":"3889","DOI":"10.1021\/acs.jmedchem.7b01729","article-title":"Combating drug-resistant fungi with novel imperfectly amphipathic palindromic peptides","volume":"61","author":"Wang","year":"2018","journal-title":"J. Med. Chem."},{"key":"B43","doi-asserted-by":"publisher","first-page":"6434","DOI":"10.1016\/j.psj.2020.09.030","article-title":"Avian host defense cathelicidins: structure, expression, biological functions, and potential therapeutic applications","volume":"99","author":"Wang","year":"2020","journal-title":"Poultry Sci."},{"key":"B44","doi-asserted-by":"publisher","first-page":"3546","DOI":"10.1021\/jm4004158","article-title":"Structure and function of a potent lipopolysaccharide-binding antimicrobial and anti-inflammatory peptide","volume":"56","author":"Wei","year":"2013","journal-title":"J. Med. Chem."},{"key":"B45","doi-asserted-by":"publisher","first-page":"2204","DOI":"10.1016\/j.biomaterials.2010.11.054","article-title":"Synthetic cationic amphiphilic \u03b1-helical peptides as antimicrobial agents","volume":"32","author":"Wiradharma","year":"2011","journal-title":"Biomaterials"},{"key":"B46","doi-asserted-by":"publisher","first-page":"74","DOI":"10.1002\/marc.201200534","article-title":"Rationally designed \u03b1-helical broad-spectrum antimicrobial peptides with idealized facial amphiphilicity","volume":"34","author":"Wiradharma","year":"2013","journal-title":"Macromol. Rapid Commun."},{"key":"B47","doi-asserted-by":"publisher","first-page":"427","DOI":"10.1016\/j.ijantimicag.2016.12.012","article-title":"and characterisation of a novel antibacterial peptide from a native swine intestinal tract-derived bacterium","volume":"49","author":"Xin","year":"2017","journal-title":"Int. J. Antimicrob. Agents"},{"key":"B48","doi-asserted-by":"publisher","first-page":"944","DOI":"10.1038\/s41598-020-58014-6","article-title":"Conversion of broad-spectrum antimicrobial peptides into species-specific antimicrobials capable of precisely targeting pathogenic bacteria","volume":"10","author":"Xu","year":"2020","journal-title":"Sci. Rep."},{"key":"B49","doi-asserted-by":"publisher","first-page":"764","DOI":"10.1002\/jsfa.11410","article-title":"Oleanolic acid reshapes the gut microbiota and alters immune-related gene expression of intestinal epithelial cells","volume":"102","author":"Xue","year":"2021","journal-title":"J. Sci. Food Agric"},{"key":"B50","doi-asserted-by":"publisher","first-page":"W174","DOI":"10.1093\/nar\/gkv342","article-title":"I-TASSER server: new development for protein structure and function predictions","volume":"43","author":"Yang","year":"2015","journal-title":"Nucleic Acids Res."},{"key":"B51","doi-asserted-by":"publisher","first-page":"10469","DOI":"10.1021\/acs.jmedchem.1c00879","article-title":"PEGylation of the antimicrobial peptide PG-1: a link between propensity for nanostructuring and capacity of the antitrypsin hydrolytic ability","volume":"64","author":"Yu","year":"2021","journal-title":"J. Med. Chem."},{"key":"B52","doi-asserted-by":"publisher","first-page":"4918","DOI":"10.1128\/AAC.00202-11","article-title":"Easy strategy to increase salt resistance of antimicrobial peptides","volume":"55","author":"Yu","year":"2011","journal-title":"Antimicrobial Agents Chemother."},{"key":"B53","doi-asserted-by":"publisher","first-page":"1307","DOI":"10.1007\/s11426-013-4884-y","article-title":"Comparison on effect of hydrophobicity on the antibacterial and antifungal activities of \u03b1-helical antimicrobial peptides","volume":"56","author":"Zhao","year":"2013","journal-title":"Sci. China Chem."},{"key":"B54","doi-asserted-by":"publisher","first-page":"244","DOI":"10.1016\/j.actbio.2013.08.043","article-title":"Design of imperfectly amphipathic \u03b1-helical antimicrobial peptides with enhanced cell selectivity","volume":"10","author":"Zhu","year":"2014","journal-title":"Acta Biomater."},{"key":"B55","doi-asserted-by":"publisher","first-page":"155","DOI":"10.1016\/j.actbio.2015.02.023","article-title":"of antimicrobial activity and mechanisms of low amphipathic peptides with different \u03b1-helical propensity","volume":"18","author":"Zhu","year":"2015","journal-title":"Acta Biomater."},{"key":"B56","doi-asserted-by":"publisher","first-page":"9421","DOI":"10.1021\/acs.jmedchem.0c00583","article-title":"Rational avoidance of protease cleavage sites and symmetrical end-tagging significantly enhances the stability and therapeutic potential of antimicrobial peptides","volume":"63","author":"Zhu","year":"2020","journal-title":"J. Med. Chem."}],"container-title":["Frontiers in Microbiology"],"original-title":[],"link":[{"URL":"https:\/\/www.frontiersin.org\/articles\/10.3389\/fmicb.2022.864374\/full","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2022,7,4]],"date-time":"2022-07-04T09:01:17Z","timestamp":1656925277000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.frontiersin.org\/articles\/10.3389\/fmicb.2022.864374\/full"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2022,7,4]]},"references-count":56,"alternative-id":["10.3389\/fmicb.2022.864374"],"URL":"https:\/\/doi.org\/10.3389\/fmicb.2022.864374","relation":{},"ISSN":["1664-302X"],"issn-type":[{"value":"1664-302X","type":"electronic"}],"subject":[],"published":{"date-parts":[[2022,7,4]]},"article-number":"864374"}}