What is substrate-level phosphorylation?

Substrate-level phosphorylation is a mechanism of generating ATP by transferring a phosphate group directly to ADP from a phosphorylated intermediate. This process occurs during specific enzymatic reactions, primarily in glycolysis and the citric acid cycle (Krebs cycle). In glycolysis, for example, a molecule of glycerate-1,3-bisphosphate donates a phosphate group to ADP to form ATP, showcasing how ATP is produced without the involvement of electron transport chains or proton gradients.

This process differs from oxidative phosphorylation, where ATP is generated indirectly through an electron transport chain and chemiosmosis. While substrate-level phosphorylation does not depend on oxygen, this characteristic is not what defines it; rather, the direct transfer of phosphate is the critical feature. Additionally, substrate-level phosphorylation is distinct from the production of NADH and FADH2, which are involved in the electron transport chain and are important for oxidative phosphorylation rather than for the direct generation of ATP in this manner.