Chemoproteomics reveals extensive cysteine S-sulfhydration in deep-sea hyperthermophilic archaea

Thermococcales are an order of archaea that inhabit the sulfur-rich, anaerobic environments of deep-sea hydrothermal vents. Their metabolism shifts toward the production of hydrogen sulfide (H₂S) in the presence of elemental sulfur. As a gaseous signaling molecule, H₂S primarily regulates biological processes by mediating the S-sulfhydration of protein cysteine. However, it remains unknown whether such modifications occur in these archaea. In this study, we employed chemical proteomics to systematically identify S-sulfhydration sites in the deep-sea hyperthermophilic archaeon Thermococcus aciditolerans SY113. This approach revealed unique reactivity characteristics of the modification and allowed quantitative analysis of its dynamic regulation by H₂S. A total of 204 S-sulfhydration sites on 171 proteins were identified, over 65% of which were dynamically regulated by H₂S. Further functional analysis indicated that S-sulfhydration plays an ancient and conserved role in primordial cells, including the regulation of catalytic activity, maintenance of protein conformation, and mediation of protein-protein interactions. Our findings provide a valuable dataset and theoretical foundation for understanding the role of S-sulfhydration in the physiological regulation of deep-sea hyperthermophilic archaea.