title: Aspartate dehydrogenase, a novel enzyme identified from structural and functional studies of TM1643.
abstract: The open reading frame TM1643 of Thermotoga maritima belongs to a large family of proteins, with homologues in bacteria, archaea, and eukaryotes. TM1643 is found in an operon with two other genes that encode enzymes involved in the biosynthesis of NAD. In several bacteria, the gene in the position occupied by TM1643 encodes an aspartate oxidase (NadB), which synthesizes iminoaspartate as a substrate for NadA, the next enzyme in the pathway. The amino acid sequence of TM1643 does not share any recognizable homology with aspartate oxidase or with other proteins of known functions or structures. To help define the biological functions of TM1643, we determined its crystal structure at 2.6A resolution and performed a series of screens for enzymatic function. The structure reveals the presence of an N-terminal Rossmann fold domain with a bound NAD(+) cofactor and a C-terminal alpha+beta domain. The structural information suggests that TM1643 may be a dehydrogenase and the active site of the enzyme is located at the interface between the two domains. The enzymatic characterization of TM1643 revealed that it possesses NAD or NADP-dependent dehydrogenase activity toward l-aspartate but no aspartate oxidase activity. The product of the aspartate dehydrogenase activity is also iminoaspartate. Therefore, our studies demonstrate that two different enzymes, an oxidase and a dehydrogenase, may have evolved to catalyze the first step of NAD biosynthesis in prokaryotes. TM1643 establishes a new class of amino acid dehydrogenases.
authors.affiliation: Department of Biological Sciences, Columbia University, New York, New York 10027, USA.
authors.forename: Zhiru
authors.initials: Z
authors.lastname: Yang
authors.forename: Alexei
authors.initials: A
authors.lastname: Savchenko
authors.forename: Alexander
authors.initials: A
authors.lastname: Yakunin
authors.forename: Rongguang
authors.initials: R
authors.lastname: Zhang
authors.forename: Aled
authors.initials: A
authors.lastname: Edwards
authors.forename: Cheryl
authors.initials: C
authors.lastname: Arrowsmith
authors.forename: Liang
authors.initials: L
authors.lastname: Tong
mesh_terms: D000594:Amino Acid Oxidoreductases / Q000737:chemistry / Q000378:metabolism*; D000595:Amino Acid Sequence; D001665:Binding Sites; D018360:Crystallography, X-Ray; D007700:Kinetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011487:Protein Conformation; D017386:Sequence Homology, Amino Acid; D013329:Structure-Activity Relationship
chemical_list: D000594:Amino Acid Oxidoreductases; C025745:aspartate dehydrogenase