Human Glutathione Synthetase (hGS) catalyzes the second step in the synthesis of the important intracellular antioxidant glutathione (gamma-glutamylcysteinylglycine): gamma-glutamylcysteine + glycine + ATP → GSH + ADP + Pi Homodimeric hGS is a member of the ATP grasp superfamily, whose members are primarily non-mammalian and involved in nucleotide biosynthesis. Human GS is much like rat GS, and while both these enzymes have similar Km values for substrates as the yeast, A. thaliana and E. coli GS, they differ in subunit numbers and in cooperativity. Only mammalian (rat and human) GS displays negative cooperativity with respect to the gamma-glutamyl substrate; glycine and ATP are not cooperative. Negative cooperativity is found in regulatory pathways where substrates of the allosteric enzyme have other functions. That human GS is negatively cooperative suggests that gamma-glutamylcysteine may have other metabolic roles/pathways of importance in addition to glutathione biosynthesis. Studies on the reaction order of the eukaryote, A. thaliana, suggested that the GS reaction is random order ternary mechanism, but not cooperative. Our current studies on human GS suggest that while glycine and ATP binding are not cooperative, they support the findings that the gamma-glutamyl substrate binding is negatively cooperative. Additionally, our current research suggests that the human reaction, unlike plant, is at least partially ordered. An understanding of human GS will increase our knowledge and assist in the development of herbicides and antibiotics that do not affect humans. Supported in part by a TWU REP grant (MEA) and a Welch Foundation grant to TWU Chemistry and Biochemistry.

Human glutathione synthetase: Negative cooperativity and substrate binding studies

255th National Meeting (2018)

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