Multiple pathways for the formation of the γ-glutamyl peptides γ-glutamyl-valine and γ- glutamyl-valyl-glycine in Saccharomyces cerevisiae

Abstract

The role of glutathione (GSH) in eukaryotic cells is well known. The biosynthesis of this γ-glutamine tripeptide is well studied. However, other γ-glutamyl peptides were found in various sources, and the pathways of their formation were not always clear. The aim of the present study was to determine whether Saccharomyces cerevisiae can produce γ-glutamyl tripeptides other than GSH and to identify the pathways associated with the formation of these peptides. The tripeptide γ-Glu-Val-Gly (γ-EVG) was used as a model. Wild-type yeast cells were shown to produce this peptide during cultivation in minimal synthetic medium. Two different biosynthetic pathways for this peptide were identified. The first pathway consisted of two steps. In the first step, γ-Glu-Val (γ-EV) was produced from glutamate and valine by the glutamate-cysteine ligase (GCL) Gsh1p or by the transfer of the γ-glutamyl group from GSH to valine by the γ-glutamyltransferase (GGT) Ecm38p or by the (Dug2p-Dug3p) 2 complex. In the next step, γ-EV was combined with glycine by the glutathione synthetase (GS) Gsh2p. The second pathway consisted of transfer of the γ-glutamyl residue from GSH to the dipeptide Val-Gly (VG). This reaction was carried out mainly by the (Dug2p-Dug3p) 2 complex, whereas the GGT Ecm38p did not participate in this reaction. The contribution of each of these two pathways to the intracellular pool of γ-EVG was dependent on cultivation conditions. In this work, we also found that Dug1p, previously identified as a Cys-Gly dipeptidase, played an essential role in the hydrolysis of the dipeptide VG in yeast cells. It was also demonstrated that γ-EV and γ-EVG could be effectively imported from the medium and that γ-EVG was imported by Opt1p, known to be a GSH importer. Our results demonstrated that γ-glutamyl peptides, particularly γ-EVG, are produced in yeast as products of several physiologically important reactions and are therefore natural components of yeast cells.

Kokumi taste perception is functional in a model carnivore, the domestic cat (Felis catus)

“…Of the potential kokumi-tasting compounds, some of the γ-glutamyl peptides, calcium, and the biogenic amine spermidine exhibited the lowest EC 50 values for cCaSR. The formation of γ-glutamyl peptides is linked to GSH metabolism, and both γ-glutamyl peptides and GSH are found in different meats 32 , 33 , meaning that these compounds are present in a carnivorous diet. Similarly, spermidine, which is a biogenic amine, is found in food sources palatable to cats, including chicken liver 42 .…”

Perturbations of the Proteome and of Secreted Metabolites in Primary Astrocytes from the hSOD1(G93A) ALS Mouse Model

“…Nonetheless, GCL presents a relatively broad substrate specificity and can bind glutamate with some other amino acids in addition to cysteine, producing the corresponding γ-glutamyl dipeptides [77]. The role of γ-glutamyl dipeptides in cells is not yet fully understood, but they have been detected in both the brain and eyes [78], indicating a potential involvement of these compounds in neuronal homeostasis.…”

“…The lower amount of GCLC and/or the lower GSH substrate availability to γ-glutamyltransferase, responsible for the transfer of the γ-glutamyl moiety to any amino acid [78], may thus account for the lower amount of the γ-glutamyl-tyrosine dipeptide detected in the CM of hSOD1(G93A) astrocytes by the metabolomic analysis.…”

Non-canonical glutamate-cysteine ligase activity protects against ferroptosis

“…For example, serine palmitoyl transferase will also metabolize alanine or glycine when serine is limiting (Penno et al, 2010) and glutamate-aspartate aminotransferase will also metabolize cysteine sulfinic acid (Weinstein et al, 1988). In the case of GCLC, this feature may have been selected for during evolution, as the S. cerevisiae homolog (Gsh1p) also has the ability to at least use valine (Sofyanovich et al, 2019). Additional work is needed to determine which other amino acids are accepted by S.…”

“…However, 13 C5, 15 N2-glutamine tracing demonstrated that while the newly labeled GSH fraction was very small, as expected, glutamate and γ-glutamyl-dipeptides were newly labeled to ~ 50% in cystine starved A549 cells ( Figure 4C and D), suggesting that the γ-glutamyl dipeptides were synthesized from glutamate but not from GSH ( Figure S4B). Because γ-glutamyl-valine is synthesized by the Saccharomyces cerevisiae glutamate-cysteine ligase (Sofyanovich et al, 2019), and γ-glutamyl dipeptide synthesis by mouse liver extracts was recently shown to be GCLC-dependent (Kobayashi et al, 2020), we hypothesized that the γ-glutamyl dipeptides were directly generated by GCLC rather than GSH metabolism by GGT. To evaluate this, we evaluated the γ-glutamyl dipeptide levels in the GCLC and GSS KO clones.…”