Nicotinamide mononucleotide synthetase is the key enzyme for an alternative route of NAD biosynthesis in Francisella tularensis

Leonardo Sorci, Dariusz Martynowski, Dmitry A. Rodionov, Yvonne Eyobo, Xhavit Zogaj, Karl E. Klose, Evgeni V. Nikolaev, Giulio Magni, Hong Zhang, Andrei L. Osterman

Research output: Contribution to journalArticle

37 Scopus citations

Abstract

Enzymes involved in the last 2 steps of nicotinamide adenine dinucleotide (NAD) cofactor biosynthesis, which catalyze the adenylylation of the nicotinic acid mononucleotide (NaMN) precursor to nicotinic acid dinucleotide (NaAD) followed by its amidation to NAD, constitute promising drug targets for the development of new antibiotics. These enzymes, NaMN adenylyltransferase (gene nadD) and NAD synthetase (gene nadE), respectively, are indispensable and conserved in nearly all bacterial pathogens. However, a comparative genome analysis of Francisella tularensis allowed us to predict the existence of an alternative route of NAD synthesis in this category A priority pathogen, the causative agent of tularaemia. In this route, the amidation of NaMN to nicotinamide mononucleotide (NMN) occurs before the adenylylation reaction, which converts this alternative intermediate to the NAD cofactor. The first step is catalyzed by NMN synthetase, which was identified and characterized in this study. A crystal structure of this enzyme, a divergent member of the NadE family, was solved at 1.9-Å resolution in complex with reaction products, providing a rationale for its unusual substrate preference for NaMN over NaAD. The second step is performed by NMN adenylyltransferase of the NadM family. Here, we report validation of the predicted route (NaMN → NMN → NAD) in F. tularensis including mathematical modeling, in vitro reconstitution, and in vivo metabolite analysis in comparison with a canonical route (NaMN → NaAD → NAD) of NAD biosynthesis as represented by another deadly bacterial pathogen, Bacillus anthracis.

Original languageEnglish (US)
Pages (from-to)3083-3088
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume106
Issue number9
DOIs
Publication statusPublished - Mar 3 2009

    Fingerprint

Keywords

  • Genomic reconstruction
  • In vitro reconstitution
  • Mathematical modeling
  • NAD intermediates
  • Substrate preference

ASJC Scopus subject areas

  • General

Cite this