Glutathione S-transferases (GSTs) are a multigene family (EC 2.5.1.18) of eight dimeric enzymes that are classified based on their amino acid sequences and substrate specificity as alpha (A), kappa (K), mu (M), omega (O), pi (P), sigma (S), theta (T), and zeta (Z) [1]. Depending on their subcellular location, GSTs are grouped as cytoplasmic (A, P, M, S, T, Z), mitochondrial (K), or membrane-bound (Membrane Associated Proteins in Eicosanoid and Glutathione metabolism) [2]. GSTs are phase-II detoxification enzymes found in most life forms and vital for maintaining cellular homeostasis [3]. GSTs play a cytoprotective role primarily by catalyzing the conjugation reaction of reduced glutathione (GSH) and reactive electrophiles generated by cytochrome P450 metabolism to form GSH conjugates [4]. The resulting GSH conjugates are either excreted via bile or transported to the kidney where: (1) the γ-glutamyl moiety is cleaved by γ-glutamyl transpeptidase; (2) the glycine is cleaved by dipeptidase; and (3) the cysteine is N-acetylated [5].

In addition to their detoxification roles, GSTs are known for their functions in cell signaling, post-translational modification, and resistance to chemotherapeutic agents [6]. For example, the pi and mu classes of GSTs regulate the mitogen-activated protein (MAP) kinase pathway that governs cell survival and cell death signals via direct interactions with c-Jun N-terminal kinase 1 (JNK1) and apoptosis signal-regulating kinase (ASK1) [7]. Additionally, GSTs form complexes with an array of intracellular proteins for their post-translational modification [8]. For instance, protein disulfide isomerase (PDI), peroxiredoxin-VI (Prdx VI), and p53 are common substrates of GST-mediated glutathionylation [9]. Similar to the detoxification process described above, antineoplastic drugs bound to GSH are expelled out of the cells by the membrane-bound GS-X pump, making cancer cells resistant to chemotherapy [10]. Since their discovery in 1961 in rat liver [11], GSTs have gained attention among cancer researchers. The expression of GSTs in all cell types and their abundance in aggressive cancer cells suggest that they play a key role in tumor progression and cancer pathogenicity [12]. Recent developments in the field of redox oncology have shed light on novel functions of GST proteins in cancer cells [13,14]. This review summarizes newly identified functions of GST proteins and their roles in the cellular signaling, metabolism, and survival of cancer cells.

Because GSTs are pivotal in drug metabolism, they were among the first cytosolic proteins to be structurally characterized. Porcine GST Pi 1 (GSTP1)was the first member of the family whose structure was determined [15]. Crystallographic studies have revealed that the catalytic GSTs display analogous tertiary structures and exist as homodimers in mammals (Figure 1A) [16]; however, heterodimers of a few cytosolic GSTs have been identified in plants [17]. Currently, no enzymatically active monomers of GST proteins are known [15]. Subsequent structural analyses revealed that all principal GST family members have a basic protein fold consisting of two domains: the N-terminal domain and the C-terminal domain. The GST N-terminal domain fold is similar to other cellular homeostasis and detoxification proteins such as glutathione peroxidases and glutaredoxins. The N-terminal domain constitutes approximately one-third of the protein structure and is made up of a β-α-β-α-β-β-α motif. The β-β-α motif in the N-terminal domain, also known as the G-site, is most conserved among the isoforms and provides the binding site for reduced glutathione (GSH) by recognizing the γ-glutamyl fragment of GSH (Figure 1B).

Interestingly, a proline residue, found at the N-terminal end of strand β3, is conserved among all cytosolic and mitochondrial GSTs. This proline forms hydrogen-bond interactions with the backbone amine group of the GSH-cysteinyl moiety (Figure 1C) [16,18,19]. Global characterization of sequence and structure similarity of GST proteins show two major subgroups: (1) tyrosine-type GSTs (Y-GSTs), which use tyrosine to activate GSH; and (2) S/C-GSTs, which use serine (or cysteine in case of GST Omega (GSTO)) to interact with GSH [20]. However, the C-terminal domain of GSTs constitutes the other two-thirds of the protein structure and is made up of a unique all-α-helical domain [19]. The hydrophobic substrates bind to a cleft between the N- and C-terminal domains known as the H-site. Unlike the G-site, the H-site is highly variable in shape and chemical constitution between classes [21]. This variability in H-site structure determines the substrate selectivity of various GST isozymes [22].

3. Conclusions and Future Directions