Each molecule of GADP is then oxidized by a molecule of NAD+ in the presence of GAP, forming 1,3-bisphosphoglycerate. In the next step, PGK generates a molecule of ATP while forming 3-phosphoglycerate. At this step, glycolysis has reached the break-even point: 2 molecules of ATP were consumed, and 2 new molecules have been synthesized. This step, one of the two substrate-level phosphorylation steps, requires ADP; thus, when the cell has plenty of ATP (and little ADP) this reaction does not occur. Because ATP decays relatively quickly when it is not metabolized, this is an important regulatory point in the glycolytic pathway. PGAM then forms 2-phosphoglycerate; ENO then forms phosphoenolpyruvate; and another substrate-level phosphorylation then forms a molecule of Pyr and a molecule of ATP by means of the enzyme PK. This serves as an additional regulatory step.
After the formation of F1,6bP, many of the reactions are energetically unfavorable. The only reactions that are favorable are the 2 substrate-level phosphorylation steps that result in the formation of ATP. These two reactions pull the glycolytic pathway to completion.
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