It has been reported that catechins have antibacterial, anti-toxin, and membrane-altering properties. For example, catechins have been demonstrated to inhibit the activity of the leukotoxin (LtxA) produced by <i>Aggregatibacter actinomycetemcomitans</i>, although the mechanism of this inhibition was unknown. We hypothesized that catechin-mediated changes in membrane properties inhibit bacterial toxin binding, thus leading to the reported anti-toxin properties. To test this hypothesis, six common catechins were used: (-)-catechin (C), (-)-epicatechin (EC), (-)-epigallocatechin (EGC), (-)-gallocatechin gallate (GCg), (-)-epigallocatechin gallate (EGCg) and (-)-epicatechin gallate (ECg) to correlate inhibition of LtxA activity with the catechins’ ability to alter the host membrane properties. Our results indicate that the catechin-mediated decrease in host cell membrane fluidity does not contribute to the reported anti-toxin mechanism of catechins. Even more, pre-incubation of LtxA with catechins is more effective in the prevention of cytotoxicity than pre-incubation of cells with catechins, suggesting that the catechins act on the toxin rather than the cell membrane to inhibit LtxA activity. Consistent with this finding, we found that the catechins decreased the affinity of LtxA for cholesterol, an interaction required for cytotoxicity. In each of these experiments, GCg, EGCg, and ECg were more effective than C, EC, and EGC. Binding of LtxA to cholesterol is mediated by a cholesterol-binding motif located in the C-terminal end of LtxA; we, therefore, suspected that catechins might alter the exposure of this domain by altering the toxin’s secondary structure. Using circular dichroism spectroscopy, we found that the secondary structure of LtxA is drastically changed when the toxin is treated with EGCg, but not with C, suggesting that galloylated catechins modify protein toxin structure, thus inhibiting the toxin from binding to the host cell surface.

Catechin-mediated restructuring of a bacterial toxin inhibits activity

2019 Spring National Meeting

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