Structural adaptation of an interacting non-native C-terminal helical extension revealed in the crystal structure of NAD+ synthetase from Bacillus anthracis.

Academic Article

Abstract

  • The crystal structures of NH(3)-dependent NAD+ synthetase from Bacillus anthracis as the apoenzyme (1.9 A), in complex with the natural catalytic products AMP and pyrophosphate (2.4 A) and in complex with the substrate analog adenosine 5'-(alpha,beta-methylene)triphosphate (2.0 A) have been determined. NAD+ synthetase catalyzes the last step in the biosynthesis of the vitally important cofactor NAD+. In comparison to other NAD+ synthetase crystal structures, the C-terminal His-tagged end of the apoenzyme adopts a novel helical conformation, causing significant compensatory changes in the region. The structural accommodations observed in B. anthracis NAD+ synthetase are remarkable in the absence of adverse affects on enzyme activity. They also illustrate a rare example of the influence of a non-native C-terminal His-tag extension on the structure of a native protein. In contrast to the apoenzyme, when AMP and pyrophosphate or adenosine 5'-(alpha,beta-methylene)triphosphate are bound, the C-terminus adopts a conformation that allows ATP binding and overall the structure then resembles other NAD+ synthetase structures. The structures of NAD+ synthetase complexes from B. anthracis are compared with published X-ray crystal structures of the enzyme from B. subtilis, Escherichia coli and Helicobacter pylori. These comparisons support the novel observation that P1 and P2 loop ordering is not a consequence of crystal contacts but rather a consequence of intrinsic intramolecular interactions within the ordered subunit.
  • Keywords

  • Adenosine Triphosphate, Amide Synthases, Amination, Amino Acid Sequence, Apoenzymes, Bacillus anthracis, Binding Sites, Conserved Sequence, Crystallography, X-Ray, Gene Expression, Histidine, Hydrogen-Ion Concentration, Models, Molecular, Molecular Sequence Data, Niacin, Phylogeny, Protein Structure, Quaternary, Protein Structure, Tertiary, Sequence Alignment, Substrate Specificity
  • Digital Object Identifier (doi)

    Author List

  • McDonald HM; Pruett PS; Deivanayagam C; Protasevich II; Carson WM; DeLucas LJ; Brouillette WJ; Brouillette CG
  • Start Page

  • 891
  • End Page

  • 905
  • Volume

  • 63
  • Issue

  • Pt 8