Journal of Applied Phycology

, 21:103 | Cite as

The cyanobacterial alkaloid nostocarboline: an inhibitor of acetylcholinesterase and trypsin

  • Paul G. Becher
  • Heike I. Baumann
  • Karl Gademann
  • Friedrich Jüttner


Preselected cyanobacterial strains (available from culture collections and our own isolates), belonging primarily to the heterocystous cluster, were screened for inhibitors against butyrylcholinesterase. About one-half of the extracts exhibited inhibitory activity. Nostocarboline, the responsible metabolite in Nostoc 78–12A, was studied in more detail as an acetylcholinesterase (AChE) inhibitor. The compound showed potent activity against this enzyme (IC50 = 5.3 µM), and the Michaelis-Menten kinetics indicated a non-competitive component in the inhibitory mechanism. In addition, nostocarboline turned out to be a potent inhibitor of trypsin (IC50 = 2.8 µM), and thus is the first described cyanobacterial serine protease inhibitor of an alkaloid structure. The function of nostocarboline in aquatic ecosystems and its potential as a lead compound for the development of useful therapeutic AChE inhibitors is discussed.


Screening Cyanobacteria Nostoc Serine protease Alzheimer’s disease 



This work was supported by the National Science Foundation, Bern, and Hydrobiologie-Limnologie Stiftung, Zürich.


  1. Bartolini M, Bertucci C, Cavrini V, Andrisano V (2003) Beta-amyloid aggregation induced by human acetylcholinesterase: inhibition studies. Biochem Pharmacol 65:407–416PubMedCrossRefGoogle Scholar
  2. Bartus RT, Dean RL, Beer B, Lippa AS (1982) The cholinergic hypothesis of geriatric memory dysfunction. Science 217:408–417PubMedCrossRefGoogle Scholar
  3. Baumann HI, Keller S, Wolter FE, Nicholson GJ, Jung G, Süssmuth RD, Jüttner F (2007) Planktocyclin, a cyclooctapeptide protease inhibitor produced by the freshwater cyanobacterium Planktothrix rubescens. J Nat Prod 70:1611–1615PubMedCrossRefGoogle Scholar
  4. Becher PG, Jüttner F (2005) Insecticidal compounds of the biofilm-forming cyanobacterium Fischerella sp. (ATCC 43239). Environ Toxicol 20:363–372PubMedCrossRefGoogle Scholar
  5. Becher PG, Beuchat J, Gademann K, Jüttner F (2005) Nostocarboline: isolation and synthesis of a new cholinesterase inhibitor from Nostoc 78–12A. J Nat Prod 68:1793–1795PubMedCrossRefGoogle Scholar
  6. Becher PG, Jüttner F (2006) Insecticidal activity—a new bioactive property of the cyanobacterium Fischerella. Pol J Ecol 54:653–662Google Scholar
  7. Becher PG, Keller S, Jung G, Süssmuth RD, Jüttner F (2007) Insecticidal activity of 12-epi-hapalindole J isonitrile. Phytochemistry 68:2493–2497PubMedCrossRefGoogle Scholar
  8. Bisswanger H (2000) Enzymkinetik—Theorie und Methoden. Wiley-VCH, WeinheimGoogle Scholar
  9. Blom J, Bister B, Bischoff D, Nicholson G, Jung G, Süssmuth RD, Jüttner F (2003) Oscillapeptin J, a new grazer toxin of the freshwater cyanobacterium Planktothrix rubescens. J Nat Prod 66:431–434PubMedCrossRefGoogle Scholar
  10. Blom JF, Brutsch T, Barbaras D, Bethuel Y, Locher HH, Hubschwerlen C, Gademann K (2006) Potent algicides based on the cyanobacterial alkaloid nostocarboline. Org Lett 8:737–740PubMedCrossRefGoogle Scholar
  11. Burja AM, Banaigs B, Abou-Mansour E, Burgess JG, Wright PC (2001) Marine cyanobacteria—a prolific source of natural products. Tetrahedron 57:9347–9377CrossRefGoogle Scholar
  12. Carmichael WW (1992) Cyanobacteria secondary metabolites—the cyanotoxins. J Appl Bacteriol 72:445–459PubMedGoogle Scholar
  13. Codd GA (1995) Cyanobacterial toxins: occurrence, properties and biological significance. Water Sci Technol 32:149–156CrossRefGoogle Scholar
  14. Dembitsky VM, Řezanka T (2005) Metabolites produced by nitrogen-fixing Nostoc species. Folia Microbiol 50:363–391CrossRefGoogle Scholar
  15. Ellman GL, Courtney KD, Andres V, Featherstone RM (1961) A new and rapid colorimetric determination of acetylcholinesterase activity. Biochem Pharmacol 7:88–95PubMedCrossRefGoogle Scholar
  16. Fahrney DE, Gold AM (1963) Sulfonyl fluorides as inhibitors of esterases. I. Rates of reaction with acetylcholine esterase, α-chymotrypsin, and trypsin. J Am Chem Soc 85:997–1000CrossRefGoogle Scholar
  17. Gademann K (2007) Cyanobacterial natural products for the inhibition of biofilm formation and biofouling. Chimia 61:373–377CrossRefGoogle Scholar
  18. Gearhart DA, Neafsey EJ, Collins MA (2002) Phenylethanolamine N-methyltransferase has β-carboline 2N-methyltransferase activity: hypothetical relevance to Parkinson’s disease. Neurochem Internat 40:611–620CrossRefGoogle Scholar
  19. Ghosal S, Bhattacharya SK, Mehta R (1972) Naturally occurring and synthetic β-carbolines as cholinesterase inhibitors. J Pharm Sci 61:808–810PubMedCrossRefGoogle Scholar
  20. Grau S, Baldi A, Bussani R, Tian X, Stefanescu R, Przybylski M, Richards P, Jones SA, Shridhar V, Clausen T, Ehrmann M (2005) Implications of the serine protease HtrA1 in amyloid precursor protein processing. Proc Nat Acad Sci USA 102:6021–6026PubMedCrossRefGoogle Scholar
  21. Haider S, Naithani V, Viswanathan PN, Kakkar P (2003) Cyanobacterial toxins: a growing environmental concern. Chemosphere 52:1–21PubMedCrossRefGoogle Scholar
  22. Harel M, Kryger G, Rosenberry TL, Mallender WD, Lewis T, Fletcher RJ, Guss JM, Silman I, Sussman JL (2000) Three-dimensional structures of Drosophila melanogaster acetylcholinesterase and of its complexes with two potent inhibitors. Protein Sci 9:1063–1072PubMedCrossRefGoogle Scholar
  23. Horowitz AR, Denholm I (2001) Impact of insecticides resistance mechanisms on management and strategies. In: Ishaaya I (ed) Biochemical sites of insecticide action and resistance. Springer, Berlin, pp 323–338Google Scholar
  24. Hostettmann K, Borloz A, Urbain A, Marston A (2006) Natural product inhibitors of acetylcholinesterase. Curr Org Chem 10:825–847CrossRefGoogle Scholar
  25. Houghton PJ, Ren Y, Howes M-J (2006) Acetylcholinesterase inhibitors from plants and fungi. Nat Prod Rep 23:181–199PubMedCrossRefGoogle Scholar
  26. Inestrosa NC, Alvarez A, Pérez CA, Moreno RD, Vicente M, Linker C, Casanueva OI, Soto C, Garrido J (1996) Acetylcholinesterase accelerates assembly of amyloid-β-peptides into Alzheimer’s fibrils: possible role of the peripheral site of the enzyme. Neuron 16:881–891PubMedCrossRefGoogle Scholar
  27. Jüttner F, Wu JT (2000) Evidence of allelochemical activity in subtropical cyanobacterial biofilms of Taiwan. Arch Hydrobiol 147:505–517Google Scholar
  28. Jüttner F, Wessel HP (2003) Isolation of di(hydroxymethyl)dihydroxypyrrolidine from the cyanobacterial genus Cylindrospermum that effectively inhibits digestive glucosidases of aquatic insects and crustacean grazers. J Phycol 39:26–32CrossRefGoogle Scholar
  29. Kawabata SI, Miura T, Morita T, Kato H, Fujikawa K, Iwanaga S, Takada K, Kimura T, Sakakibara S (1988) Highly sensitive peptide-4-methylcoumaryl-7-amide substrates for blood-clotting proteases and trypsin. Eur J Biochem 172:17–25PubMedCrossRefGoogle Scholar
  30. Kleinschmidt S, Ziegeler S, Bauer C (2005) Cholinesterase inhibitors. Importance in anaesthesia, intensive care medicine, emergency medicine and pain therapy. Anaesthesist 54:791–799PubMedCrossRefGoogle Scholar
  31. Kraut D, Goff H, Pai RK, Hosea NA, Silman I, Sussman JL, Taylor P, Voet JG (2000) Inactivation studies of acetylcholinesterase with phenylmethylsulfonyl fluoride. Mol Pharmacol 57:1243–1248PubMedGoogle Scholar
  32. Kuhn W, Muller T, Grosse H, Rommelspacher H (1996) Elevated levels of harman and norharman in cerebrospinal fluid of Parkinsonian patients. J Neural Transm 103:1435–1440PubMedCrossRefGoogle Scholar
  33. Liao D-I, Qian J, Chisholm DA, Jordan DB, Diner BA (2000) Crystal structures of the photosystem II D1 C-terminal processing protease. Nat Struct Biol 7:749–753PubMedCrossRefGoogle Scholar
  34. Mahmood NA, Carmichael WW (1987) Anatoxin-a(s), an anticholinesterase from the cyanobacterium Anabaena flos-aquae NRC-525–17. Toxicon 25:1221–1227PubMedCrossRefGoogle Scholar
  35. Matsubara K, Collins MA, Akane A, Ikebuchi J, Neafsey EJ, Kagawa M, Shiono H (1993) Potential bioactivated neurotoxicants, N-methylated β-carbolinium ions, are present in human brain. Brain Res 610:90–96PubMedCrossRefGoogle Scholar
  36. Metcalf JS, Codd GA (2004) Cyanobacterial toxins in the water environment. A review of current knowledge. Foundation for Water Research, Marlow, UKGoogle Scholar
  37. Miota F, Siegfried BD, Scharf ME, Lydy MJ (2000) Atrazine induction of cytochrome P450 in Chironomus tentans larvae. Chemosphere 40:285–291PubMedCrossRefGoogle Scholar
  38. Nair JS, Ramaswamy NK (2004) Chloroplast proteases. Biol Planta 48:321–326CrossRefGoogle Scholar
  39. Park H-J, Kim S-S, Seong Y-M, Kim K-H, Goo HG, Yoon EJ, Min, DS, Kang S, Rhim H (2006) β-Amyloid precursor protein is a direct cleavage target of HtrA2 serine protease. Implications for the physiological function of HtrA2 in the mitochondria. J Biol Chem 281:34277–34287PubMedCrossRefGoogle Scholar
  40. Robinson ESJ, Anderson NJ, Crosby J, Nutt DJ, Hudson AL (2003) Endogenous β-carbolines as clonidine-displacing substances. Ann N Y Acad Sci 1009:157–166PubMedCrossRefGoogle Scholar
  41. Schott Y, Decker M, Rommelspacher H, Lehmann J (2006) 6-Hydroxy- and 6-methoxy-β-carbolines as acetyl- and butyrylcholinesterase inhibitors. Bioorg Med Chem Lett 16:5840–5843PubMedCrossRefGoogle Scholar
  42. Scott LJ, Goa KL (2000) Galantamine: a review of its use in Alzheimer’s disease. Drugs 60:1095–1122PubMedCrossRefGoogle Scholar
  43. Selkoe DJ (1999) Translating cell biology into therapeutic advances in Alzheimer’s disease. Nature 399:A23–31PubMedCrossRefGoogle Scholar
  44. Sturm A, Hansen PD (1999) Altered cholinesterase and monooxygenase levels in Daphnia magna and Chironomus riparius exposed to environmental pollutants. Ecotoxicol Environ Saf 42:9–15PubMedCrossRefGoogle Scholar
  45. Sussman JL, Silman I (1992) Acetylcholinesterase: structure and use as a model for specific cation-protein interactions. Curr Opin Struct Biol 2:721–729CrossRefGoogle Scholar
  46. Taylor P (1991) The cholinesterases. J Biol Chem 266:4025–4028PubMedGoogle Scholar
  47. Teichert A, Schmidt J, Porzel A, Arnold N, Wessjohann L (2007) Brunneins A-C, β-carboline alkaloids from Cortinarius brunneus. J Nat Prod 70:1529–1531PubMedCrossRefGoogle Scholar
  48. Todorova AK, Jüttner F, Linden A, Plüss T, Philipsborn WV (1995) Nostocyclamide: a new macrocyclic, thiazole-containing allelochemical from Nostoc sp. 31 (Cyanobacteria). J Org Chem 60:7891–7895CrossRefGoogle Scholar
  49. Trost JT, Chisholm DA, Jordan DB, Diner BA (1997) The D1 C-terminal processing protease processing of photosystem II from Scenedesmus obliquus. Protein purification and gene characterization in wild type and processing mutants. J Biol Chem 272:20348–20356PubMedCrossRefGoogle Scholar
  50. Volk RB (2005) Screening of microalgal culture media for the presence of algicidal compounds and isolation and identification of two bioactive metabolites, excreted by the cyanobacteria Nostoc insulare and Nodularia harveyana. J Appl Phycol 17:339–347CrossRefGoogle Scholar
  51. Volk RB, Furkert FH (2006) Antialgal, antibacterial and antifungal activity of two metabolites produced and excreted by cyanobacteria during growth. Microbiol Res 161:180–186PubMedCrossRefGoogle Scholar
  52. Volk RB (2007) Studies on culture age versus exometabolite production in batch cultures of the cyanobacterium Nostoc insulare. J Appl Phycol 19:491–495CrossRefGoogle Scholar
  53. Volk RB, Mundt S (2007) Cytotoxic and non-cytotoxic exometabolites of the cyanobacterium Nostoc insulare. J Appl Phycol 19:55–62CrossRefGoogle Scholar
  54. Welker M, von Döhren H (2006) Cyanobacterial peptides—Nature’s own combinatorial biosynthesis. FEMS Microbiol Rev 30:530–563PubMedCrossRefGoogle Scholar
  55. Wylie CR, Paul VJ (1988) Feeding preferences of the surgeonfish Zebrasoma flavescens in relation to chemical defenses of tropical algae. Mar Ecol Prog Ser 45:23–32CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media B.V. 2008

Authors and Affiliations

  • Paul G. Becher
    • 1
  • Heike I. Baumann
    • 1
  • Karl Gademann
    • 2
  • Friedrich Jüttner
    • 1
  1. 1.Limnological Station, Institute of Plant BiologyUniversity of ZürichKilchbergSwitzerland
  2. 2.Chemical Synthesis Laboratory, SB-ISIC-LSYNCSwiss Federal Institute of Technology (EPFL)LausanneSwitzerland

Personalised recommendations