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Abstract
The metabolic network of Saccharomyces cerevisiae is highly flexible in response to rapid changes in the environment. Amino acid metabolism plays an important role in many cellular processes and this thesis addresses dynamic changes in amino acid metabolism during several physiological changes including: heat shock, one-carbon metabolism and anaerobiosis.
Glycine and L-serine are interconvertible one-carbon donors, while L-threonine is the major supplier of glycine in cells. Previous work demonstrated that glycine addition led to induction of the one-carbon regulon, and L-serine addition led to the biosynthesis of pyruvate. Interestingly, L-threonine addition resulted in the shuttling of L-threonine to L-serine and subsequent catabolism to pyruvate. Transcriptional data showed that L-threonine-derived glycine did not induce the one-carbon regulon; instead it induced the glycine decarboxylase/serine hydroxymethyltransferase system and catabolic L-serine (L-threonine) deaminase to enable the continuous flow from L-threonine to pyruvate. The addition of L-threonine may have activated the GAAC response as a result of an imbalance in metabolism. This response is mediated by Gcn4p, the master regulator of amino acid biosynthesis.
Gcn4p and L-serine also play important roles in anaerobic amino acid metabolism. The lack of L-serine in anaerobic gcn4 cells induced an extended lag phase as a result of L-serine starvation. The availability of L-serine was crucial for the biosynthesis of serine-rich cell wall mannoproteins that are essential and specific to anaerobic cells. Mutation of GCN4 led to the inability of cells to biosynthesise L-serine from L-threonine, thus relying on biosynthesis from 3-phosphoglycerate and/ or the glyoxylate route.
L-Tryptophan has been previously shown to be involved in ageing and heat shock resistance. The addition of L-tryptophan to cells led to increased synthesis of L-kynurenine and nicotinate, which were found to be metabolites that assist cells in heat tolerance (50°C). The addition of L-tryptophan induced a gene expression pattern that has some similarity to the response observed in heat shocked cells, thus the addition of L-tryptophan could have possibly activated the transcription of genes required for heat stress. This thesis has demonstrated the importance of amino acid metabolism in a variety of cellular processes.