Phenylalanine dehydrogenase from Rhodococcus sp. M4: High-resolution X-rayanalyses of inhibitory ternary complexes reveal key features in the oxidative deamination mechanism

The molecular structures of recombinant L-phenylalanine dehydrogenase from Rhodococcus sp. M4 in two different inhibitory ternary complexes have been determined by X-ray crystallographic analyses to high resolution. Both stru ctures show that L-phenylalanine dehydrogenase is a homodimeric enzyme with each monomer composed of distinct globular N- and C-terminal domains separ ated by a deep cleft containing the active site. The N-terminal domain bind s the amino acid substrate and contributes to the interactions at the subun it:subunit interface. The C-terminal domain contains a typical Rossmann fol d and orients the dinucleotide. The dimer has overall dimensions of approxi mately 82 Angstrom x 75 Angstrom x 75 Angstrom, with roughly 50 Angstrom se parating the two active sites. The structures described here, namely the en zyme NAD(+).phenylpyruvate, and enzyme NAD(+).beta-phenylpropionate species , represent the first models for any amino acid dehydrogenase in a ternary complex. By analysis of the active-site interactions in these models, along with the currently available kinetic data, a detailed chemical mechanism h as been proposed. This mechanism differs from those proposed to date in tha t it accounts for the inability of the amino acid dehydrogenases, in genera l, to function as hydroxy acid dehydrogenases.