Phosphorylation occurs at serine 113. Isocitrate dehydrogenase is inactivated by phosphorylation through an effect on substrate binding. Phosphorylation may prevent an induced conformational change necessary for substrate binding. The dephosphorylated form is found in a pocket containing numerous positively charged and other polar residues. Inactivation of isocitrate dehydrogenase by phosphorylation occurs by preventing isocitrate from binding. The inhibition of binding is caused by the introduction of a negative charge at position 113. The phosphorylation sits is in a pocket lined with polar side chains. Structural changes are restricted to the region of the phosphorylation site; no protein moves more that 0.4Å. The lack of a large-scale conformational change in the unliganded enzyme suggests isocitrate dehydrogenase is inactivated by electrostatic isocitrate-phosphoserine interaction (9). The serine that is phosphorylated may be needed to bind NADP+ or for catalysis. The phosphate may mask other residues or sterically block a binding site. The conformation of the enzyme may be altered by the addition of the large phosphate or negative charge (11). The phosphorylation system efficiently compensates for varying isocitrate dehydrogenase levels by regulating the phosphorylation state of isocitrate dehydrogenase, resulting in a nearly constant isocitrate dehydrogenase activity during growth of E. coli on acetate. (10)

       The replacement of serine with aspartate results in inactivation of isocitrate dehydrogenase due to the negative charge of Asp. Substitution of other amino acids with larger side chains, but less charge impacts, like tyrosine, result in a decrease in activity, but not complete inactivation. The substitutions showed an increase in Km, but no change in Vmax of the enzyme (11).