title: Facilitative production of an antimicrobial peptide royalisin and its antibody via an artificial oil-body system.
abstract: Royalisin found in the royal jelly of Apis mellifera is an antimicrobial peptide (AMP). It has a molecular weight of 5.5 kDa, which contains six cysteine residues. In this study, royalisin was overexpressed in Escherichia coli AD494 (DE3) as two oleosin-fusion proteins for preparation of its antibodies and functional purification. The recombinant royalisin, fused with oleosin central hydrophobic domain in both N- and C-termini, was reconstituted with triacylglycerol and phospholipids to form artificial oil bodies (AOBs). The AOBs were then purified to raise the antibodies. These antibodies could recognize both the native and recombinant royalisins, but not oleosin. Another oleosin-intein S-fusion protein was purified by AOBs system, and royalisin was subsequently released from the AOBs through self-splicing of the intein. The recombinant royalisin exhibited high antibacterial activity, which suggested that it was refolded to its functional structure. These results demonstrated that AOBs system is an efficient method to functionally express and purify small AMPs. In addition, it also provides a facile platform for the production of antibodies against small peptides.
authors.affiliation: Graduate Institute of Biotechnology, National Chung-Hsing University, Taichung, Taiwan.
authors.forename: Jun-Ming
authors.initials: JM
authors.lastname: Tseng
authors.forename: Jun-Ru
authors.initials: JR
authors.lastname: Huang
authors.forename: Hsiou-Chen
authors.initials: HC
authors.lastname: Huang
authors.forename: Jason T C
authors.initials: JT
authors.lastname: Tzen
authors.forename: Wing-Ming
authors.initials: WM
authors.lastname: Chou
authors.forename: Chi-Chung
authors.initials: CC
authors.lastname: Peng
mesh_terms: D000818:Animals; D000900:Anti-Bacterial Agents / Q000378:metabolism* / Q000494:pharmacology; D000906:Antibodies / Q000276:immunology*; D000935:Antifungal Agents / Q000378:metabolism* / Q000494:pharmacology; D001483:Base Sequence; D001516:Bees; D015153:Blotting, Western; D017931:DNA Primers; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D036341:Intercellular Signaling Peptides and Proteins; D008826:Microbial Sensitivity Tests; D010957:Plasmids; D011506:Proteins / Q000235:genetics* / Q000276:immunology / Q000494:pharmacology; D011994:Recombinant Proteins / Q000235:genetics
chemical_list: D000900:Anti-Bacterial Agents; D000906:Antibodies; D000935:Antifungal Agents; D017931:DNA Primers; D036341:Intercellular Signaling Peptides and Proteins; D011506:Proteins; D011994:Recombinant Proteins; C064722:royalisin