Pseudomonas aeruginosa is an important human pathogen associated with several acute and
chronic conditions, including diseases of the airways and
wounds. The organism produces
pyocyanin, an extracellular redox toxin that induces oxidative stress, depletes intracellular
glutathione (GSH) and induces proliferative arrest and apoptosis, thus compromising the ability of tissue to repair itself. GSH is an important intra- and extracellular
antioxidant, redox
buffer and detoxifies
xenobiotics by increasing their polarity, which facilitates their elimination. As previous studies have reported exogenous GSH to be protective against
pyocyanin toxicity, this study was undertaken to explore the mechanism by which GSH protects host cells from the deleterious effects of the toxin. Co-incubation of
pyocyanin with GSH resulted in a time-dependent diminished recovery of the toxin from the incubation medium. Concurrently, a highly polar green-colored metabolite was recovered that exhibited a UV-visible spectrum similar to
pyocyanin and which was determined by mass spectrometry to have a major ion (m/z = 516) consistent with a
glutathione conjugate. The ability of the conjugate to oxidize
NADPH and to reduce molecular
oxygen with the production of
reactive oxygen species was comparable to
pyocyanin yet it no longer demonstrated cytotoxicity towards host cells. These data suggest that GSH forms a cell-impermeant conjugate with
pyocyanin and that availability of the
thiol may be critical to minimizing the toxicity of this important bacterial
virulence factor at
infection sites. Our data indicate that for GSH to have a clinically effective role in neutralizing
pyocyanin, the
thiol needs to be available at millimolar concentrations.