Enhancing cellular persulfides through artificial substrate of 3 mercaptopyruvate sulfurtansferase 3 mst

Abstract

Hydrogen sulfide (H2S) is a major gaseous signalling molecule that is responsible for redox regulation within cells, having implications for primary metabolism, antibiotic response, health, and ageing. 3-Mercaptopyruvate sulfurtransferase (3-MST) is one of the key enzyme involved in the biosynthesis of H2S which occurs through a crucial intermediate step called as 3-MST persulfidation (MST-SSH). Persulfide generated from the 3-MST pathway is essential for regulating mitochondrial activity and transporting sulfur via downstream sulfide transfer to proteins, low molecular weight thiols, and Fe-S clusters. Precise mechanisms by which these effects are mediated by 3-MST remain poorly understood. Also, several studies have shown that dysfunction or dysregulation of 3-MST is linked to various pathophysiological conditions. Hence, the development of a reliable tool is required in order to have better understanding of 3-MST activity as well as 3-MST expression level. In order to understand the biological effects of 3-MST, we hypothesized to modulate the substrate delivery to the enzyme. Owing to the limitations associated with the native substrate (3-mercaptopyruvate), it can not be used reliably to enhance 3-MST activity. In this regard, our lab identified 2-mercapto-1-phenylethan-1-one (PhCOCH2SH) as an artificial substrate for 3-MST that produces H2S/persulfide in cells. Following this, a library of PhCOCH2SH analogues was synthesized to tune the rate of H2S and persulfide production. Structure-activity study with analogues revealed that 3-MST was fairly accommodative of structural modifications. Linear free energy relationship study showed that H2S release rates from para-substituted aryl substrates in the presence of 3-MST had a moderate positive slope supporting a transition state with a no major charge build up. Ortho substituents slowed H2S generation rates likely due to the steric effects and molecular docking studies showed that this substitution pattern reduced access to the active site due to steric clash