Biologically relevant hydrogen sulfide chemistry: Redox interaction with thiol proteins

A new paradigm in signal transduction was established with the discovery that the simple diatomic nitric oxide (NO) is endogenously generated as a crucial and fundamental signaling molecule. This discovery spurred the search and study of other endogenously produced small molecules that, like nitric oxide, could regulate biological functions. Two other molecules, carbon monoxide (CO) and hydrogen sulfide (H2S), are now known to be signaling molecules and along with nitric oxide have been termed "gasotransmitters". (It should to be noted, however, that these molecules are solutes at the concentrations that elicit biological activity. Thus, the unfortunate term "gasotransmitters" refers to the physical state of the pure species at room temperature and pressure and not to their state in biological systems.) These molecules have become the subjects of intense and recent investigations focused on understand their biological roles and delineating their signaling properties/biochemistry. In the case of hydrogen sulfide, this molecule has been known for hundreds of years as a toxic gas, but in recent times production in mammalian cells has been established. A variety of studies have alluded to important physiological functions of this small signaling molecule. However, the chemical/biochemical basis for these activities remains obscure. The work presented here is aimed at determining whether H2S can participate in biological thiol redox chemistry. It is hypothesized that H2S can interact with oxidized cysteine moieties of proteins resulting in the production of perthiol intermediates that have either novel biological activity or serve as a precursor to eventual generation of the fully reduced cysteine. To this end, a study of the ability of H2S to restore the activity of oxidized glyceraldehyde-3-phosphate dehydrogenase (GAPDH, a classic cysteine-based dehydrogenase) and papain (a cysteine-based protease) by reducing their oxidized thiolate active site has been performed. Our results indicate that H2S is capable of reducing disulfide and sulfenic acid forms of GAPDH. Moreover, using an oxidized glutathione model system to examine the chemical mechanism by which H2S reacts with oxidized thiol species it is found that reduction of a disulfide bond by H2S leads to formation of a stable perthiol intermediate. These results support a mechanism in which reduction of oxidized GAPDH and papain by hydrogen sulfide involves a perthiol intermediate. Based on these results a hypothesis is forwarded that biological perthiol formation is facile, the perthiol intermediate is fairly stable and represents a relevant and important aspect of thiol redox biochemistry.