Crystal structure of NH3-dependent NAD+ synthetase from Bacillus subtilis

EMBO J. 1996 Oct 1;15(19):5125-34.


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.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amide Synthases*
  • Ammonia
  • Bacillus subtilis / enzymology*
  • Binding Sites
  • Crystallography, X-Ray
  • Dimerization
  • Ligases / chemistry*
  • Models, Chemical
  • Models, Molecular
  • NAD / biosynthesis*
  • Protein Conformation*
  • Recombinant Proteins / chemistry


  • Recombinant Proteins
  • NAD
  • Ammonia
  • Ligases
  • Amide Synthases
  • NAD+ synthase