## 3 Results and Discussion
### Cellular Amino Acid Metabolic Process
#### 3.3.1 Cellular Amino Acid Metabolic Process at the Middle of the Second Fermentation  
At the middle of the second fermentation, 24 specific proteins in the P29 strain and 28 specific proteins in the G1 strain were identified. These proteins are represented as nodes in Figure 2A,B, respectively. A total of 28 interactions (number of edges) were observed, with a \(p\)-value of PPI enrichment <2.23 \(\times\) 10\({}^{-12}\) in the P29 strain; while in the G1 strain, 36 interactions were obtained, with a \(p\)-value of PPI enrichment <4.44 \(\times\) 10\({}^{-16}\); such enrichment indicates that the proteins are at least partially biologically connected as a group. In the P29 strain, the strongest interactions were observed between proteins belonging to lysine metabolism (white nodes) and the metabolism of branched chain amino acids (dark blue nodes). These amino acids are used for the synthesis of numerous volatile  
Figure 2: High confidence protein–protein interaction network map built using STRING v11 and based on the proteins of cellular amino acid metabolic process detected in _S. cerevisiae_ P29 (**A**) and G1 (**B**) in the middle of the second fermentation. Proteins are shown as nodes, and the existence of interactions between them are represented by lines (the connection between nodes). Line thickness indicates the strength of the different interactions. Nodes with the same color represent specific clusters: sulfur amino acid metabolic process (red nodes), branched-chain amino acid metabolic process (light blue nodes), glutamine metabolic process (light green nodes), glycine metabolic process (yellow nodes), and glutamate metabolic process (light purple nodes). aromatic compounds, such as higher alcohols. Furthermore, interactions were obtained between proteins belonging to different GO terms: Gdh3p-Aat1p, Uga3p-Dal81p, and Put1p-Uga2p. The Snz3p, Xbp1p, and Ths1p proteins showed no interaction with the rest of the proteins (Figure 2A). On the other hand, the strongest interactions present in the G1 strain were between proteins belonging to the metabolism of sulfur amino acids, the metabolism of lysine, and the metabolism of glycine. However, most proteins are involved with the metabolism of sulfur amino acids such as methionine and cysteine, which are necessary for yeast growth [33]. Lap3p, Sno3p, and Ehd3p did not show any interaction but were included in different GO terms (Figure 2B). In both strains, the metabolism of lysine was identified. This could be related to an antioxidant response by yeasts [34]. The main difference between both strains in the middle of the second fermentation is glutamate metabolism in the P29 strain and glycine and tryptophan metabolism in the G1 strain. These data are in agreement with the results obtained by [PERSON] et al. (2016) [6]. These authors reported an increase in the assimilation of various amino acids, particularly glutamate and glycine. Both amino acids are related to the synthesis of most amino acids. Furthermore, the reversible conversion of glutamate to \(\alpha\)-ketoglutarate is an important branch point between carbon and nitrogen metabolism [6]. In addition, glutamate is one of the nitrogen sources that maintain a high growth rate [35]. During the early stages of fermentation, _S. cerevisiae_ rapidly accumulates those amino acids necessary for protein synthesis and formation of volatile compounds such as higher alcohols and their respective ethyl and acetic esters [12], while the surplus is stored in the cellular vacuole, because an amino acid deficit causes a decrease in cell biomass [21]. During the fermentation process, bioactive compounds such as melatonin and serotonin may form because of the tryptophan metabolism, in addition to the higher alcohol tryptophan, which are compounds that have an impact on the sensory properties of the wine [22]. In this case, the identified protein belonging to the tryptophan metabolism, Bna4p (0.01 mol\(\%\)), is a kynureine 3-monooxygenase.