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Title: Mechanism of action of recombinant Acc-royalisin from royal jelly of Chinese honeybee against gram-positive bacteria

Author
item SHEN, LIRONG - Zhejiang University
item LIU, DANDAN - Zhejiang University
item YANG, XIA0LI - Zhejiang University
item JIN, FENG - Zhejiang University
item DIN, MEIHUI - Zhejiang University
item Parnell, Laurence
item Lai, Chao Qiang

Submitted to: PLOS ONE
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 9/11/2012
Publication Date: 10/9/2012
Citation: Shen, L., Liu, D., Yang, X., Jin, F., Din, M., Parnell, L.D., Lai, C. 2012. Mechanism of action of recombinant Acc-royalisin from royal jelly of Chinese honeybee against gram-positive bacteria. PLoS One. 7(10):e47194 DOI: 10.1371/journal.pone.0047194.

Interpretive Summary: The royal jelly produced by honeybees is a very popular supplement in Asian countries for its potential benefits in human health. One key function of royal jelly is its antibacterial activity of royalisin, an antimicrobial peptide. Due to its safety and effectiveness, royalisin has a great potential application for food preservation, therapeutic application and medication. In this study, we isolated and purified the recombinant royalisin expressed from the E. coli BL21, which we cloned previously from Chinese honeybee. We further demonstrated that the expressed and purified royalisin inhibits the growth of Gram-positive bacteria, similar to nisin, a polycyclic antibacterial peptide currently used as a food preservative. The antibacterial activity of the royalisin was associated with its concentration, and was weakened by heat treatment at 50 deg C and 80 deg C for 15 min. Using transmission electron microscopy, our results further illustrated that royalisin inhibits the growth of bacteria likely by inducing the disruption and dysfunction of cell walls and membranes. This is the first study detailing the antibacterial mechanism of royalisin against Gram-positive bacteria, and provides insight into the application of recombinant royalisin in food and pharmaceutical industries as an antimicrobial agent.

Technical Abstract: The antibacterial activity of royalisin, an antimicrobial peptide from the royal jelly produced by honeybees has been addressed extensively. However, its mechanism of action remains unclear. In this study, a recombinant royalisin, RAcc-royalisin from the royal jelly of Chinese honeybee Apis cerana cerana, was expressed fused with glutathione S-transferase (GST) in Escherichia coli DL21, isolated and purified. The agar dilution assays with inhibition zone showed that RAcc-royalisin inhibits the growth of Gram-positive bacteria, similar to nisin. The antibacterial activity of RAcc-royalisin was associated with its concentration, and was weakened by heat treatment at 50 deg C and 80 deg C for 15 min. RAcc-royalisin exhibited the minimum inhibitory concentrations (MIC) of 25 ug/ml and 50 ug/ml against B. subtilis and M. flavus in the microplate assay, respectively. The antibacterial activity of RAcc-royalisin at these MIC levels agrees well with the decrease in bacterial cell hydrophobicity, the leakage of 260-nm absorbing materials, and the observation by transmission electron microscopy, all indicating that RAcc-royalisin induced the disruption and dysfunction of cell walls and membranes. This is the first report detailing the antibacterial mechanism of royalisin against Gram-positive bacteria, and provides insight into the application of recombinant royalisin in food and pharmaceutical industries as an antimicrobial agent.