Crystal structure of NH3-dependent NAD+ synthetase from Bacillus subtilis.
NAD+ synthetase catalyzes the last step in the biosynthesis of nicotinamide adenine dinucleotide. The three-dimensional structure of NH3-dependent NAD+ synthetase from Bacillus subtilis, in its free form and in complex with ATP, has been solved by X-ray crystallography (at 2.6 and 2.0 angstroms resolution, respectively) using a combination of multiple isomorphous replacement and density modification techniques. The enzyme consists of a tight homodimer with alpha/beta subunit topology. The catalytic site is located at the parallel beta-sheet topological switch point, where one AMP molecule, one pyrophosphate and one Mg2+ ion are observed. Residue Ser46, part of the neighboring 'P-loop', is hydrogen bonded to the pyrophosphate group, and may play a role in promoting the adenylation of deamido-NAD+ during the first step of the catalyzed reaction. The deamido-NAD+ binding site, located at the subunit interface, is occupied by one ATP molecule, pointing towards the catalytic center. A conserved structural fingerprint of the catalytic site, comprising Ser46, is very reminiscent of a related protein region observed in glutamine-dependent GMP synthetase, supporting the hypothesis that NAD+ synthetase belongs to the newly discovered family of 'N-type' ATP pyrophosphatases.
Department of Genetics and Microbiology A. Buzzati Traverso, Universityof Pavia, Italy.
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Mattevi
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Bolognesi
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Galizzi
D019732:Amide Synthases; D000641:Ammonia; D001412:Bacillus subtilis / Q000201:enzymology*; D001665:Binding Sites; D018360:Crystallography, X-Ray; D019281:Dimerization; D008025:Ligases / Q000737:chemistry*; D008956:Models, Chemical; D008958:Models, Molecular; D009243:NAD / Q000096:biosynthesis*; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry
D011994:Recombinant Proteins; D009243:NAD; D000641:Ammonia; D008025:Ligases; D019732:Amide Synthases; C038479:NAD+ synthase