Plasmodium falciparum causes the deadliest form of malaria. Adequate redox control is crucial for this protozoan parasite to overcome oxidative and nitrosative challenges, thus enabling its survival. Sulfenylation is an oxidative post-translational modification, which acts as a molecular on/off switch, regulating protein activity. To obtain a better understanding of which proteins are redox regulated in malaria parasites, we established an optimized affinity capture protocol coupled with mass spectrometry analysis for identification of in vivo sulfenylated proteins. The non-dimedone based probe BCN-Bio1 shows reaction rates over 100-times that of commonly used dimedone-based probes, allowing for a rapid trapping of sulfenylated proteins. Mass spectrometry analysis of BCN-Bio1 labeled proteins revealed the first insight into the Plasmodium falciparum trophozoite sulfenylome, identifying 102 proteins containing 152 sulfenylation sites. Comparison with Plasmodium proteins modified by S-glutathionylation and S-nitrosation showed a high overlap, suggesting a common core of proteins undergoing redox regulation by multiple mechanisms. Furthermore, parasite proteins which were identified as targets for sulfenylation were also identified as being sulfenylated in other organisms, especially proteins of the glycolytic cycle. This study suggests that a number of Plasmodium proteins are subject to redox regulation and it provides a basis for further investigations into the exact structural and biochemical basis of regulation, and a deeper understanding of cross-talk between post-translational modifications.
Keywords: 9-hydroxymethylbicyclo[6.1.0]nonyne (BCN-Bio1); Cysteine modification; Malaria; Post-translational modification; Redox proteomics; Sulfenylation.
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