Involvement of Pseudomonas aeruginosa Rhodanese in Protection from Cyanide Toxicity

Abstract

Cyanide is a serious environmental pollutant and a biocontrol metabolite in plant growth-promoting Pseudomonas species. Here we report on the presence of multiple sulfurtransferases in the cyanogenic bacterium Pseudomonas aeruginosa PAO1 and investigate in detail RhdA, a thiosulfate:cyanide sulfurtransferase (rhodanese) which converts cyanide to less toxic thiocyanate. RhdA is a cytoplasmic enzyme acting as the principal rhodanese in P. aeruginosa. The rhdA gene forms a transcriptional unit with the PA4955 and psd genes and is controlled by two promoters located upstream of PA4955 and rhdA. Both promoters direct constitutive RhdA expression and show similar patterns of activity, involving moderate down-regulation at the stationary phase or in the presence of exogenous cyanide. We previously observed that RhdA overproduction protects Escherichia coli against cyanide toxicity, and here we show that physiological RhdA levels contribute to P. aeruginosa survival under cyanogenic conditions. The growth of a ⌬rhdA mutant is impaired under cyanogenic conditions and fully restored upon complementation with rhdA. Wild-type P. aeruginosa outcompetes the ⌬rhdA mutant in cyanogenic coculture assays. Hence, RhdA could be regarded as an effector of P. aeruginosa intrinsic resistance to cyanide, insofar as it provides the bacterium with a defense mechanism against endogenous cyanide toxicity, in addition to cyanide-resistant respiration.

Repurposing the antimycotic drug flucytosine for suppression ofPseudomonas aeruginosapathogenicity

“…The PpvdE::lux construct was integrated into the genome of P. aeruginosa strains as described (30). The PAO1pvdS mutant was generated by replacement of the entire pvdS coding sequence with a Gm R cassette using a previously described strategy (31). The inframe deletion mutants PAO1codA and PAO1codB were generated using the suicide vector pDM4 as described (32).…”

Common themes and variations in the rhodanese superfamily

“…The capacity of Bacillus stearothermophilus to detoxify cyanide is greatly increased in mutants showing 5-to 6-folds rhodanese activity than wild-type (54). Moreover, RhdA from the cyanogenic bacterium P. aeruginosa has recently been reported to contribute to cyanide detoxification (8,14,53). In fact, although RhdA is endowed with low affinity for both thiosulfate and cyanide in vitro, it has been shown to provide protection against cyanide toxicity when overexpressed in E. coli as heterologous host (53).…”

“…Accordingly, here we present the rhodanese-like proteins characterized so far grouped into four distinct classes, likely corresponding to homogeneous structural groups. Interestingly, genome analysis of P. aeruginosa reveals the presence of ten rhodanese-like proteins (14) belonging to each of the rhodanese groups here proposed, with the only exception of the members belonging to the elongated active-site loop proteins group (i.e., group IV), a class of rhodanese-like proteins that are apparently restricted to eukaryotic organisms. Thus, the coexistence of several rhodanese-like proteins in the same organism suggests a plethora of different physiological roles fulfilled by the rhodanese superfamily.…”

Rhodanese Functions as Sulfur Supplier for Key Enzymes in Sulfur Energy Metabolism

“…Despite numerous studies, the physiological role of rhodaneses as well as their physiological substrates remain unclear and are still widely debated. It has been proposed that they accomplish essential cell functions as they may be involved in the maintenance of redox homeostasis (20), in the elimination of toxic cyanide (21), and in the biosynthesis of several other cellular metabolites such as vitamins, enzymes, and cofactors that include a step of sulfur transfer (22)(23)(24)(25).…”