May 3, 2002
Department of Food Science
Initial Ph.D. Seminar
Olive-Jean Burrowes
Thesis Advisor: Dr. Tung-Ching Lee
Production of Pleurocidin in Genetically Engineered Yeast
for use in Food Applications
The emergence of antimicrobial resistant organisms in food is of major concern
globally. Traditionally, chemical preservatives play a major role in food
preservation, however, there is increasing consumer awareness regarding their
long-term deleterious effect on human health and the environment. Natural
antimicrobial peptides produced by microorganisms, animal and marine species,
have potential applications as food preservatives because they destroy microbial
cells by rapidly disrupting their membranes. However, continued research is
required to determine their appropriateness for use in foods.
Pleurocidin is a 25 amino acid antimicrobial peptide isolated from the winter
flounder (Pleuronectes americanus). Previous studies demonstrated that
pleurocidin is effective against both Gram positive and Gram negative bacteria, is
salt tolerant and heat stable. Amidated pleurocidin also significantly reduced the
mortality level of coho salmon infected with Vibrio anguillarum, a common fish
pathogen in aquaculture. Pleurocidin is presently chemically synthesized at
laboratory scale levels and has not been tested for its efficacy against foodborne
microorganisms or for its effect on human cells.
The objective of this research is to evaluate the suitability of pleurocidin as a
potential food preservative and to develop a system of production from
genetically modified organisms.
Foodborne microorganisms were tested against pleurocidin, using a liquid
based minimal inhibitory concentration technique. The hemolytic and cytotoxic
effects on human erythrocytes and intestinal cells were also investigated, using
colorimetric analysis of cells at varying concentrations of pleurocidin. We have
also transformed the high-yielding methylotrophic yeast Pichia pastoris, with the
pleurocidin gene. Pleurocidin was found to be active against foodborne bacteria
including Vibrio alginolyticus and against yeasts and molds, without deleterious
effects on human cells.
Our findings are of microbiological significance in food safety, aquaculture and
in medicine. Vibrio species against which pleurocidin is effective are implicated in
foodborne infections from seafood and are prominent pathogens in aquaculture.
Antimicrobial peptides are known to function in the primary immune response of
their hosts. Pleurocidin could, therefore, potentially be used as a natural food
preservative, as a therapeutic agent in aquaculture and for medicinal purposes.
References:
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Cole, A., Weis, P., Diamond, G.1997. Isolation and Characterization of Pleurocidin, an Antimicrobial Peptide in
the Skin Secretions of Winter Flounder. Journal of Biological Chemistry, 272:12008-12013.
Hancock, R., and Lehrer, R. 1998. Cationic Peptides: a new source of antibiotics. Trends in Biotechnology.
16:82-87
Jia, X., Patryzkat, A., Delvin, R., Ackerman, P., Iwama, G., Hancock, R. 2000. Antimicrobial Peptides Protect
Coho Salmon from Vibrio Anguillarum infections. Appl. and Env. Microbiol. 66: 1928-32.
Tossi, A., Sandri, l., Giangaspero, A. 2000. Amphipathic, -helical Antimicrobial Peptides. Biopolymers. 55:430.