Glutamate 270 plays an essential role in K+‐activation and domain closure of Thermus thermophilus isopropylmalate dehydrogenase

摘要

ThemutantE270AofThermusthermophilus3‐isopropylmalatedehydrogenaseexhibitslargelyreduced(～1%)catalyticactivityandnegligibleactivationbyK+comparedtothewild‐typeenzyme.A3–4kcal/molincreaseintheactivationenergyofthecatalysedreactionuponthismutationcouldalsobepredictedbyQM/MMcalculations.IntheX‐raystructureoftheE270AmutantawatermoleculewasobservedtotaketheplaceofK+.SAXSandFRETexperimentsrevealedtheessentialroleofE270instabilisationoftheactivedomain‐closedconformationoftheenzyme.Inaddition,E270seemstopositionK+intocloseproximityofthenicotinamideringofNAD+andtheelectron‐withdrawingeffectofK+mayhelptopolarisethearomaticringinordertoaidthehydride‐transfer...