Structural basis for an atypical active site of an l -aspartate/glutamate-specific racemase from Escherichia coli

We determined the crystal structure of EcL-DER to elucidate protein function and substrate specificity. Unlike other asp/glu racemases, EcL-DER has an unbalanced pair of catalytic residues, Thr83/Cys197, at the active site that is crucial for l- to d-unidirectional racemase activity. EcL-DER exhibited racemase activity for both l-glutamate and l-aspartate, but had threefold higher activity for l-glutamate. Based on the structure of the EcL-DER^C197S mutant in complex with l-glutamate, we determined the binding mode of the l-glutamate substrate in EcL-DER and provide a structural basis for how the protein utilizes l-glutamate as a main substrate. The unidirectionality, despite an equilibrium constant of unity, can be understood in terms of the Haldane relationship. We determined a crystal structure of l-specific asp/glu racemases from E. coli (EcL-DER). Atypical active site of EcL-DER for racemerization of l-specific amino acid was elucidated. Substrate binding mode of l-glutamate to EcL-DER was revealed.