amide synthases


Summary: Enzymes that catalyze the joining of either ammonia or an amide with another molecule, in which the linkage is in the form of a carbon-nitrogen bond. EC 6.3.1.

Top Publications

  1. McDonald H, Pruett P, Deivanayagam C, Protasevich I, Carson W, Delucas L, et al. Structural adaptation of an interacting non-native C-terminal helical extension revealed in the crystal structure of NAD+ synthetase from Bacillus anthracis. Acta Crystallogr D Biol Crystallogr. 2007;63:891-905 pubmed
    ..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. ..
  2. Moro W, Yang Z, Kane T, Brouillette C, Brouillette W. Virtual screening to identify lead inhibitors for bacterial NAD synthetase (NADs). Bioorg Med Chem Lett. 2009;19:2001-5 pubmed publisher
    ..Over 200 commercial compounds were purchased and evaluated in enzyme inhibition and antibacterial assays. 18 compounds inhibited NADs at or below 100 microM (7.6% hit rate), and two were selected for future SAR studies. ..
  3. Ariyanayagam M, Oza S, Guther M, Fairlamb A. Phenotypic analysis of trypanothione synthetase knockdown in the African trypanosome. Biochem J. 2005;391:425-32 pubmed
    ..Escape mutants arose after 2 weeks of induction, with all parameters, including growth, returning to normal. Selective inhibitors of TryS are required to fully validate this novel drug target. ..
  4. Hara N, Yamada K, Terashima M, Osago H, Shimoyama M, Tsuchiya M. Molecular identification of human glutamine- and ammonia-dependent NAD synthetases. Carbon-nitrogen hydrolase domain confers glutamine dependency. J Biol Chem. 2003;278:10914-21 pubmed
    ..Our molecular identification of NAD synthetases may prove useful to learn more of mechanisms regulating cellular NAD metabolism. ..
  5. Jauch R, Humm A, Huber R, Wahl M. Structures of Escherichia coli NAD synthetase with substrates and products reveal mechanistic rearrangements. J Biol Chem. 2005;280:15131-40 pubmed
    ..Phylogenetic structure comparisons suggest that the present results are relevant for designing species-specific antibiotics. ..
  6. Flohe L. The trypanothione system and the opportunities it offers to create drugs for the neglected kinetoplast diseases. Biotechnol Adv. 2012;30:294-301 pubmed publisher
    ..Heterologous expression, functional characterization and crystallization of recombinant system components finally enable structure-based rational inhibitor design...
  7. Rizzi M, Bolognesi M, Coda A. A novel deamido-NAD+-binding site revealed by the trapped NAD-adenylate intermediate in the NAD+ synthetase structure. Structure. 1998;6:1129-40 pubmed
    ..Moreover, the structural data suggest that product release can take place only after all substrates are bound to the enzyme, and product release is ultimately controlled by the conformation adopted by two mobile loops. ..
  8. Yamaguchi F, Koga S, Yoshioka I, Takahashi M, Sakuraba H, Ohshima T. Stable ammonia-specific NAD synthetase from Bacillus stearothermophilus: purification, characterization, gene cloning, and applications. Biosci Biotechnol Biochem. 2002;66:2052-9 pubmed
    ..coli was 180-fold that of B. stearothermophilus H-804. The specific assay of ammonia and ATP (up to 25 microM) with this stable NAD synthetase was possible. ..
  9. Comini M, Dirdjaja N, Kaschel M, Krauth Siegel R. Preparative enzymatic synthesis of trypanothione and trypanothione analogues. Int J Parasitol. 2009;39:1059-62 pubmed publisher
    ..The protocol also allows the synthesis of related glutathione conjugates. It will greatly facilitate the thorough analysis of this parasite's metabolism and drug screening approaches against trypanothione-dependent enzymes...

More Information


  1. Comini M, Guerrero S, Haile S, Menge U, Lünsdorf H, Flohe L. Validation of Trypanosoma brucei trypanothione synthetase as drug target. Free Radic Biol Med. 2004;36:1289-302 pubmed
  2. Oza S, Wyllie S, Fairlamb A. Mapping the functional synthetase domain of trypanothione synthetase from Leishmania major. Mol Biochem Parasitol. 2006;149:117-20 pubmed
  3. Lin C, Kwon D, Bollinger J, Walsh C. Evidence for a glutathionyl-enzyme intermediate in the amidase activity of the bifunctional glutathionylspermidine synthetase/amidase from Escherichia coli. Biochemistry. 1997;36:14930-8 pubmed
  4. Suda Y, Tachikawa H, Yokota A, Nakanishi H, Yamashita N, Miura Y, et al. Saccharomyces cerevisiae QNS1 codes for NAD(+) synthetase that is functionally conserved in mammals. Yeast. 2003;20:995-1005 pubmed
    ..Finally, the rat homologue of QNS1 was cloned and shown to functionally replace yeast QNS1, indicating that NAD(+) synthetase is functionally conserved from bacteria to yeast and mammals. ..
  5. Fyfe P, Oza S, Fairlamb A, Hunter W. Leishmania trypanothione synthetase-amidase structure reveals a basis for regulation of conflicting synthetic and hydrolytic activities. J Biol Chem. 2008;283:17672-80 pubmed publisher
    ..The potential inhibitory role of the C terminus provides a mechanism to control relative levels of the critical metabolites, trypanothione, glutathionylspermidine, and spermidine in Leishmania. ..
  6. Bieganowski P, Pace H, Brenner C. Eukaryotic NAD+ synthetase Qns1 contains an essential, obligate intramolecular thiol glutamine amidotransferase domain related to nitrilase. J Biol Chem. 2003;278:33049-55 pubmed
  7. Velu S, Mou L, Luan C, Yang Z, Delucas L, Brouillette C, et al. Antibacterial nicotinamide adenine dinucleotide synthetase inhibitors: amide- and ether-linked tethered dimers with alpha-amino acid end groups. J Med Chem. 2007;50:2612-21 pubmed
    ..Studies on nonspecific effects, including detergent properties and promiscuous inhibition, suggested little contribution to observed activities. ..
  8. Comini M, Menge U, Wissing J, Flohe L. Trypanothione synthesis in crithidia revisited. J Biol Chem. 2005;280:6850-60 pubmed
  9. Kang G, Kim Y, Im Y, Rho S, Lee J, Eom S. Crystal structure of NH3-dependent NAD+ synthetase from Helicobacter pylori. Proteins. 2005;58:985-8 pubmed publisher
  10. Amssoms K, Oza S, Ravaschino E, Yamani A, Lambeir A, Rajan P, et al. Glutathione-like tripeptides as inhibitors of glutathionylspermidine synthetase. Part 1: Substitution of the glycine carboxylic acid group. Bioorg Med Chem Lett. 2002;12:2553-6 pubmed
    ..The boronic acid appears the most promising lead compound (IC(50) of 17.2 microM). ..
  11. Oza S, Tetaud E, Ariyanayagam M, Warnon S, Fairlamb A. A single enzyme catalyses formation of Trypanothione from glutathione and spermidine in Trypanosoma cruzi. J Biol Chem. 2002;277:35853-61 pubmed
    ..The recombinant enzyme possesses weak amidase activity and can hydrolyze trypanothione, homotrypanothione, or glutathionylspermidine to glutathione and the corresponding polyamine...
  12. Chen S, Lin C, Kwon D, Walsh C, Coward J. Design, synthesis, and biochemical evaluation of phosphonate and phosphonamidate analogs of glutathionylspermidine as inhibitors of glutathionylspermidine synthetase/amidase from Escherichia coli. J Med Chem. 1997;40:3842-50 pubmed
    ..88 microM. A non-polyamine-containing phosphonamidate exhibits no significant inhibition of the synthetase or amidase activity. ..
  13. Nessi C, Albertini A, Speranza M, Galizzi A. The outB gene of Bacillus subtilis codes for NAD synthetase. J Biol Chem. 1995;270:6181-5 pubmed
    ..The time course of synthesis of OutB showed that synthesis of the enzyme started during germination and outgrowth, and reached the highest level at the end of exponential growth. The enzyme could be recovered from dormant spores. ..
  14. Pai C, Chiang B, Ko T, Chou C, Chong C, Yen F, et al. Dual binding sites for translocation catalysis by Escherichia coli glutathionylspermidine synthetase. EMBO J. 2006;25:5970-82 pubmed
    ..Our results constitute the first structural information on the biochemical features of parasite homologs (including TryS) that underlie their broad specificity for polyamines. ..
  15. Laronde Leblanc N, Resto M, Gerratana B. Regulation of active site coupling in glutamine-dependent NAD(+) synthetase. Nat Struct Mol Biol. 2009;16:421-9 pubmed publisher
  16. Oza S, Ariyanayagam M, Aitcheson N, Fairlamb A. Properties of trypanothione synthetase from Trypanosoma brucei. Mol Biochem Parasitol. 2003;131:25-33 pubmed
    ..Thus, the biosynthesis and degradation of trypanothione are similar in African and American trypanosomes, and different from the insect trypanosomatid, C. fasciculata. ..
  17. Veiga Malta I, Duarte M, Dinis M, Madureira P, Ferreira P, Videira A. Identification of NAD+ synthetase from Streptococcus sobrinus as a B-cell-stimulatory protein. J Bacteriol. 2004;186:419-26 pubmed
    ..Stimulation with the recombinant NAD(+) synthetase was also observed with other B-cell markers, such as CD19(+), B220(+), and CD21(+). Cell proliferation follows the activation induced by the recombinant NAD(+) synthetase...
  18. Amssoms K, Oza S, Augustyns K, Yamani A, Lambeir A, Bal G, et al. Glutathione-like tripeptides as inhibitors of glutathionylspermidine synthetase. Part 2: substitution of the glycine part. Bioorg Med Chem Lett. 2002;12:2703-5 pubmed
    ..Compounds with basic side chains such as diaminopropionic acid were found to be good inhibitors (K(i): 7.2 microM). Substitution of the glycine part abolished the GspS substrate properties of the tripeptide. ..
  19. Gerdes S, Kurnasov O, Shatalin K, Polanuyer B, Sloutsky R, Vonstein V, et al. Comparative genomics of NAD biosynthesis in cyanobacteria. J Bacteriol. 2006;188:3012-23 pubmed
    ..Positional clustering and the co-occurrence profile of the respective genes across a diverse collection of cellular organisms provide evidence of horizontal transfer events in the evolutionary history of this pathway. ..
  20. Kwon D, Lin C, Chen S, Coward J, Walsh C, Bollinger J. Dissection of glutathionylspermidine synthetase/amidase from Escherichia coli into autonomously folding and functional synthetase and amidase domains. J Biol Chem. 1997;272:2429-36 pubmed
  21. Wojcik M, Seidle H, Bieganowski P, Brenner C. Glutamine-dependent NAD+ synthetase. How a two-domain, three-substrate enzyme avoids waste. J Biol Chem. 2006;281:33395-402 pubmed
    ..Six distinct classes of Qns1 mutants that fall within the glutaminase domain and the synthetase domain selectively inhibit components of the coordinated reaction. ..
  22. Oza S, Chen S, Wyllie S, Coward J, Fairlamb A. ATP-dependent ligases in trypanothione biosynthesis--kinetics of catalysis and inhibition by phosphinic acid pseudopeptides. FEBS J. 2008;275:5408-21 pubmed publisher
    ..This phosphinate analogue remains the most potent enzyme inhibitor identified to date, and represents a good starting point for drug discovery for trypanosomiasis and leishmaniasis. ..
  23. Lin C, Chen S, Kwon D, Coward J, Walsh C. Aldehyde and phosphinate analogs of glutathione and glutathionylspermidine: potent, selective binding inhibitors of the E. coli bifunctional glutathionylspermidine synthetase/amidase. Chem Biol. 1997;4:859-66 pubmed
    ..In E. coli, the Gsp synthetase is part of a bifunctional enzyme opposed by the hydrolytic Gsp amidase...
  24. Resto M, Yaffe J, Gerratana B. An ancestral glutamine-dependent NAD(+) synthetase revealed by poor kinetic synergism. Biochim Biophys Acta. 2009;1794:1648-53 pubmed publisher
    ..Furthermore, the data here reported in combination with structural data available for glutamine-dependent NAD(+) synthetase lays the foundation for further investigation on the mechanism of active site coupling in these enzymes...
  25. Devedjiev Y, Symersky J, Singh R, Jedrzejas M, Brouillette C, Brouillette W, et al. Stabilization of active-site loops in NH3-dependent NAD+ synthetase from Bacillus subtilis. Acta Crystallogr D Biol Crystallogr. 2001;57:806-12 pubmed
    ..A second and novel Mg(2+) position has been observed closer to the NaAD-binding site in the structure crystallized at pH 7.5, where the enzyme is active. This could therefore be the catalytically active Mg(2+). ..
  26. Oza S, Ariyanayagam M, Fairlamb A. Characterization of recombinant glutathionylspermidine synthetase/amidase from Crithidia fasciculata. Biochem J. 2002;364:679-86 pubmed
    ..GspS can also hydrolyse trypanothione at about 1.5% of the rate with glutathionylspermidine. A single amino acid mutation (Cys-79-->Ala) is shown to ablate the amidase activity without affecting the synthetase activity...
  27. Symersky J, Devedjiev Y, Moore K, Brouillette C, DeLucas L. NH3-dependent NAD+ synthetase from Bacillus subtilis at 1 A resolution. Acta Crystallogr D Biol Crystallogr. 2002;58:1138-46 pubmed
    ..A mechanism is proposed for the second catalytic step, which includes a nucleophilic attack by the ammonia molecule on the intermediate. ..
  28. Olin Sandoval V, González Chávez Z, Berzunza Cruz M, Martínez I, Jasso Chávez R, Becker I, et al. Drug target validation of the trypanothione pathway enzymes through metabolic modelling. FEBS J. 2012;279:1811-33 pubmed publisher
    ..In contrast, use of highly potent and specific inhibitors for TryR and the antioxidant machinery is necessary to affect the antioxidant capabilities of the parasites...
  29. Bollinger J, Kwon D, Huisman G, Kolter R, Walsh C. Glutathionylspermidine metabolism in Escherichia coli. Purification, cloning, overproduction, and characterization of a bifunctional glutathionylspermidine synthetase/amidase. J Biol Chem. 1995;270:14031-41 pubmed
    ..and Tabor, C. W. (1975) J. Biol. Chem. 250, 2648-2654). Trends in Km and kcat for a set of difluorosubstituted spermidine derivatives suggest that the enzyme may bind the minor, deprotonated form of the amine nucleophile...
  30. Koch O, Cappel D, Nocker M, Jäger T, Flohe L, Sotriffer C, et al. Molecular dynamics reveal binding mode of glutathionylspermidine by trypanothione synthetase. PLoS ONE. 2013;8:e56788 pubmed publisher
    ..By inhibitor docking, the binding site for N(8)-glutathionylspermidine was characterised as druggable. ..
  31. Willison J, Tissot G. The Escherichia coli efg gene and the Rhodobacter capsulatus adgA gene code for NH3-dependent NAD synthetase. J Bacteriol. 1994;176:3400-2 pubmed
    ..coli gene. In accordance with the nomenclature proposed for Salmonella typhimurium (K. T. Hughes, B. M. Olivera, and J. R. Roth, J. Bacteriol. 170:2113-2120, 1988), the efg and adgA genes should now be designated nadE...
  32. Bieganowski P, Brenner C. The reported human NADsyn2 is ammonia-dependent NAD synthetase from a pseudomonad. J Biol Chem. 2003;278:33056-9 pubmed
    ..Here we establish that the so-called NADsyn2 is simply ammonia-dependent NAD+ synthetase from Pseudomonas, which is encoded on an operon with nicotinic acid phosphoribosyltransferase and, in some Pseudomonads, with nicotinamidase. ..
  33. Oza S, Shaw M, Wyllie S, Fairlamb A. Trypanothione biosynthesis in Leishmania major. Mol Biochem Parasitol. 2005;139:107-16 pubmed
    ..fasciculata. It also appears that the L. major still harbours a redundant GSPS pseudogene that may be currently in the process of being lost from its genome...
  34. Rizzi M, Nessi C, Mattevi A, Coda A, Bolognesi M, Galizzi A. Crystal structure of NH3-dependent NAD+ synthetase from Bacillus subtilis. EMBO J. 1996;15:5125-34 pubmed
  35. Wyllie S, Oza S, Patterson S, Spinks D, Thompson S, Fairlamb A. Dissecting the essentiality of the bifunctional trypanothione synthetase-amidase in Trypanosoma brucei using chemical and genetic methods. Mol Microbiol. 2009;74:529-40 pubmed publisher
    ..The synthetase function of TRYS is thus unequivocally validated as a drug target by both chemical and genetic methods...
  36. Torrie L, Wyllie S, Spinks D, Oza S, Thompson S, Harrison J, et al. Chemical validation of trypanothione synthetase: a potential drug target for human trypanosomiasis. J Biol Chem. 2009;284:36137-45 pubmed publisher
    ..Taken together, these data provide initial chemical validation of TryS as a drug target in T. brucei. ..
  37. Galm U, Dessoy M, Schmidt J, Wessjohann L, Heide L. In vitro and in vivo production of new aminocoumarins by a combined biochemical, genetic, and synthetic approach. Chem Biol. 2004;11:173-83 pubmed
    ..This resulted in the formation of 32 new aminocoumarin compounds. The structures of these compounds were elucidated using FAB-MS and (1)H-NMR spectroscopy. ..
  38. Deng W, Wang S, Chen Q, Zhang Z, Hu X. Effect of salt treatment on theanine biosynthesis in Camellia sinensis seedlings. Plant Physiol Biochem. 2012;56:35-40 pubmed publisher
    ..Together, these data revealed that theanine synthesis takes place both in root and shoot and CsTS accumulation is positively affected by salt treatment. ..
  39. Burbaeva G, Boksha I, Sudakov S, Miasoedov S, Savushkina O, Tereshkina E, et al. [The complex neurochemical assessment of brain proteins in mentally healthy subjects and schizophrenic patients]. Zh Nevrol Psikhiatr Im S S Korsakova. 2008;108:44-50 pubmed
    ..Thus, we suppose that mentally healthy controls and patients with schizophrenia are objectively divided into different "metabolic types". ..
  40. Begley T, Kinsland C, Mehl R, Osterman A, Dorrestein P. The biosynthesis of nicotinamide adenine dinucleotides in bacteria. Vitam Horm. 2001;61:103-19 pubmed
    ..Adenylation of this mononucleotide followed by amide formation completes the biosynthesis of NAD. An additional phosphorylation gives NADP. This review focuses on the mechanistic enzymology of this pathway in bacteria. ..
  41. Pacholec M, Freel Meyers C, Oberthür M, Kahne D, Walsh C. Characterization of the aminocoumarin ligase SimL from the simocyclinone pathway and tandem incubation with NovM,P,N from the novobiocin pathway. Biochemistry. 2005;44:4949-56 pubmed
  42. Keating T, Marshall C, Walsh C. Vibriobactin biosynthesis in Vibrio cholerae: VibH is an amide synthase homologous to nonribosomal peptide synthetase condensation domains. Biochemistry. 2000;39:15513-21 pubmed
  43. Foster M, Forrester M, Stamler J. A protein microarray-based analysis of S-nitrosylation. Proc Natl Acad Sci U S A. 2009;106:18948-53 pubmed publisher
  44. Mobley J, Poliakov A. Detection of early unfolding events in a dimeric protein by amide proton exchange and native electrospray mass spectrometry. Protein Sci. 2009;18:1620-7 pubmed publisher
  45. Bellinzoni M, Buroni S, Pasca M, Guglierame P, Arcesi F, De Rossi E, et al. Glutamine amidotransferase activity of NAD+ synthetase from Mycobacterium tuberculosis depends on an amino-terminal nitrilase domain. Res Microbiol. 2005;156:173-7 pubmed
    ..This domain, a new type of glutamine amide transfer (GAT) domain, is the first to be characterized in bacterial NAD(+) synthetases. ..
  46. Rizzi M, Nessi C, Bolognesi M, Coda A, Galizzi A. Crystallization of NAD+ synthetase from Bacillus subtilis. Proteins. 1996;26:236-8 pubmed
    ..and related bacteria. ..
  47. Savage H, Montoya G, Svensson C, Schwenn J, Sinning I. Crystal structure of phosphoadenylyl sulphate (PAPS) reductase: a new family of adenine nucleotide alpha hydrolases. Structure. 1997;5:895-906 pubmed
    ..The open, reduced form of PAPS reductase is able to bind PAPS, whereas the closed oxidized form cannot. A movement between the two monomers of the dimer may allow this switch in conformation to occur. ..
  48. Tran A, Andersson B, Pettersson U, Aslund L. Trypanothione synthetase locus in Trypanosoma cruzi CL Brener strain shows an extensive allelic divergence. Acta Trop. 2003;87:269-78 pubmed
    ..Such allelic divergence observed in T. cruzi genes might have implications for drug design against Chagas' disease and the evolutional impact of the CL Brener strain. ..
  49. Ajay A, Srivastava D. Microtubular conductometric biosensor for ethanol detection. Biosens Bioelectron. 2007;23:281-4 pubmed
    ..The sensor shows minor interference with other functional groups and alcohols. The possible causes for such interference have been discussed. ..
  50. Micheli V, Sestini S, Rocchigiani M, Jacomelli G, Manzoni F, Peruzzi L, et al. Hypoxanthine-guanine phosphoribosyltransferase deficiency and erythrocyte synthesis of pyridine coenzymes. Life Sci. 1999;64:2479-87 pubmed
    ..These findings suggest that raised NAD concentrations in HPRT- erythrocytes are due to enhanced synthesis as a result of increased enzyme activities...
  51. Pai C, Wu H, Lin C, Wang A. Structure and mechanism of Escherichia coli glutathionylspermidine amidase belonging to the family of cysteine; histidine-dependent amidohydrolases/peptidases. Protein Sci. 2011;20:557-66 pubmed publisher
    ..The structural results presented here not only elucidate the catalytic mechanism and regulation of GspA but also help us to design small molecules to inhibit or probe for the activity of GspA. ..
  52. Mueller E, Palenchar P. Using genomic information to investigate the function of ThiI, an enzyme shared between thiamin and 4-thiouridine biosynthesis. Protein Sci. 1999;8:2424-7 pubmed
  53. Markus M, Doliveira L, Malakian K, Keeney D, Severin A, Underwood K, et al. 1H, 13C, and 15N backbone assignments and secondary structure for the 60.8 kD dimer of the NAD+ synthetase from Bacillus subtilis. J Biomol NMR. 2004;28:301-2 pubmed