We performed metabolome and metabolite-metabolite correlation analyses for eight single-gene deletion mutants of Saccharomyces cerevisiae to evaluate the physiology of glucose metabolism. The irreversible enzyme reactions can become bottlenecks when intracellular metabolism is perturbed by direct interference from the central metabolic pathway by gene deletions or by a deletion of transcriptional regulator. Metabolome data reveal that transcriptional factor, gcr2, regulates the reaction that converts 3-phosphoglycerate into phosphoenolpyruvate. Metabolome data also suggest that the reaction catalyzed by pyruvate kinase makes one of the rate-limiting reactions throughout the glycolytic pathway.