title: Design, characterization and expression of a novel hybrid peptides melittin (1-13)-LL37 (17-30).
abstract: Hybridizing of different antimicrobial peptides (AMPs) has been a common practice for obtaining novel hybrid AMPs with elevated antibacterial activity but minimized cytotoxicity. The hybrid peptides melittin (1-13)-LL37 (17-30) (M-L) combining the hydrophobic N-terminal fragment of melittin (M) with the core antibacterial fragment of LL37 (L), was designed for the first time to explore its antibacterial activity and hemolytic activity against bacteria and sheep erythrocyte respectively. Results showed that M-L had an even more potent antibacterial activity against all indicator strains (especially gram-positive bacteria) than M and L, whereas didn't exhibit hemolytic activity to sheep erythrocytes, implying M-L can be served as a potential therapeutic drug to substitute traditional antibiotics. However the high expense of biosynthesis limited its further research, therefore fusion expression of M-L was carried out in Escherichia coli (E. coli) for overproducing the hybrid peptide so as to solve the problem. The DNA sequence encoding M-L with preferred codons was cloned into the pET-SUMO vector for protein expression in E. coli BL21 (DE3). After IPTG induction, approximately 165 mg soluble fusion protein SUMO-M-L was recovered per liter supernatant of the fermentation ultrasonic lysate using Ni-NTA Sepharose column (92 % purity). And 23 mg recombinant M-L was obtained per liter culture after cleavage of SUMO protease and purification of Ni-NTA Sepharose column. In sum, this research not only supplied an effective approach for overproducing hybrid peptide M-L, but paved the way for its further exploration on pharmaceutical potential and medical importance.
authors.affiliation: Laboratory of Feed Biotechnology, State Key Laboratory of Animal Nutrition, College of Animal Science & Technology, China Agricultural University, No. 2 Yuanmingyuan West Road, Haidian District, Beijing, 100193, People's Republic of China.
authors.forename: Rujuan
authors.initials: R
authors.lastname: Wu
authors.forename: Qing
authors.initials: Q
authors.lastname: Wang
authors.forename: Zhaojun
authors.initials: Z
authors.lastname: Zheng
authors.forename: Longmei
authors.initials: L
authors.lastname: Zhao
authors.forename: Yajing
authors.initials: Y
authors.lastname: Shang
authors.forename: Xubiao
authors.initials: X
authors.lastname: Wei
authors.forename: Xiudong
authors.initials: X
authors.lastname: Liao
authors.forename: Rijun
authors.initials: R
authors.lastname: Zhang
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000900:Anti-Bacterial Agents / Q000096:biosynthesis* / Q000737:chemistry / Q000494:pharmacology; D023181:Antimicrobial Cationic Peptides; D001516:Bees; D054804:Cathelicidins / Q000737:chemistry* / Q000235:genetics / Q000494:pharmacology; D002478:Cells, Cultured; D015195:Drug Design; D004912:Erythrocytes / Q000166:cytology / Q000187:drug effects; D004926:Escherichia coli / Q000187:drug effects / Q000235:genetics / Q000254:growth & development; D015870:Gene Expression; D006461:Hemolysis / Q000187:drug effects; D006801:Humans; D008089:Listeria monocytogenes / Q000187:drug effects / Q000254:growth & development; D008555:Melitten / Q000737:chemistry* / Q000235:genetics / Q000494:pharmacology; D016982:Micrococcus luteus / Q000187:drug effects / Q000254:growth & development; D008969:Molecular Sequence Data; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000494:pharmacology; D012756:Sheep; D013211:Staphylococcus aureus / Q000187:drug effects / Q000254:growth & development
chemical_list: D000900:Anti-Bacterial Agents; D023181:Antimicrobial Cationic Peptides; D054804:Cathelicidins; D011993:Recombinant Fusion Proteins; D008555:Melitten