This work aims to investigate the role of the loopβ3αC amino acids in the structure and function of isocitrate dehydrogenase kinase/phosphatase(AceK).The results demonstrate that the precise configuration of loopβ3αC is very important for AceK structure and function: structural changes alter the affinity of the enzyme for the isocitrate dehydrogenase(ICDH),which modifies enzyme activity.Intriguingly,D340 is significant for the retention of kinase and phosphatase activities,for the conformational stability of AceK,and for binding ICDH.The deletion Δ341-345 increases enzyme activity by increasing the maximum velocity and affinity for ICDH.The β3αC loop is thus critical for the structure and function of AceK.The information can contribute to understand more fully the regulatory mechanisms of AceK.