Publisher Summary The chapter discusses the biologically relevant chemistry of glutathione (GSH) and its occurrence in microbial cells. The GSH-related biochemical reactions and the physiological roles of GSH are summarized. The biosynthesis of GSH is remarkable in two ways: it is mRNA independent, and the glutamic residue is joined in an unusual peptide linkage of the γ -carbon atom to the cysteine residue. Due to this structural peculiarity, GSH is protected against proteolytic cleavage. The GSH status of cells is defined by the total concentration of GSH and the nature and distribution of the possible forms of occurrence of the tripeptide in the cell. GSH and related compounds are widespread in the microbial world, especially amongst organisms with an aerobic lifestyle. This observation emphasizes the role of GSH in cellular protection against by-products generated by oxidative metabolism, but it does not limit its functions to this role. Glutathione acts as an enzyme cofactor, transport component, nucleophilic substrate, and sulphur reservoir; and participates in key cellular processes such as protein synthesis and degradation, regulation of enzyme activity, synthesis of DNA, and maintenance of the integrity of cell membranes and organelles. Having a functional diversity, GSH is interrelated with a number of metabolic pathways and its intracellular modulation could have an impact on the entire cell, making it extremely difficult to associate directly a given cellular end-point with one molecule or system. Both in Escherichia coli and Saccharomyces cerevisiae, GSH plays an important role in cellular protection during chemical stresses in spite of the fact that key enzymes of detoxification, such as GSH peroxidase and GSH S-transferase, remain at a low level.