Abstract
This paper describes the purification and characterization of microviridin J, a newly discovered metabolite of Microcystis that causes a lethal molting disruption in Daphnia spp., upon ingestion of living cyanobacterial cells. Microviridin J consists of an acetylated chain of 13 amino acids arranged in three rings and two side chains. Unlike other known isoforms of microviridin, microviridin J contains arginine that imparts a unique solution conformation characterized by proximal hydrophobic interactions between Arg and other regions of the molecule. This eventually results in the formation and stabilization of an additional ring system. Microviridin J potently inhibits porcine trypsin, bovine chymotrypsin, and daphnid trypsin-like proteases. The activity against trypsin is most likely due to Arg and its distinctive conformational interactions. Overall, the data presented for microviridin J emphasize once again the ability of cyanobacteria to produce numerous and potent environmental toxins.
Similar content being viewed by others
References
Agrawal, M. K., Bagchi, D., and Bagchi, S. N. 2001. Acute inhibition of protease and suppression of growth in zooplankter, Moina macrocopa, by Microcystis blooms collected in Central India. Hydrobiologia 464:37–44.
Allinger, N. 1997. MM2 (QCPE Quantum Exchange program).
Bartels, C., Xia, T. H., Billeter, M., Güntert, P., and Wüthrich, K. 1995. The program XEASY for computer-supported NMR spectral analysis of biological macromolecules. J. Biomol. NMR 6:1–10.
Bieth, J., Spiess, B., and Wermuth, C. G. 1974. The synthesis and analytical use of a highly sensitive and convenient substrate of elastase. Biochem. Med. 11:350–357.
Bradford, M. 1976. A rapid and sensitive method for the quantification of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem. 72:248–254.
Braunschweiler, L. and Ernst, R. R. 1983. Coherence transfer by isotropic mixing: Application to proton correlation spectroscopy. J. Magn. Reson. 53:521–528.
Carmichael, W. W. 1992. Cyanobacteria secondary metabolites—The cyanotoxins. J. Appl. Bacteriol. 72:445–459.
Chorus, I. and Bartram, J. 1999. Toxic cyanobacteria in water. A guide to their public health consequences, monitoring and management. World Health Organization.
Christoffersen, K. 1996. Ecological implications of cyanobacterial toxins in aquatic food webs. Phycologia 35:42–50.
Codd, G. A. 1995. Cyanobacterial toxins: Occurrence, properties and biological significance. Water Sci. Technol. 32:149–156.
DeMott, W. R. and Dhawale, S. 1995. Inhibition of in vitro protein phosphatase activity in three zooplankton species by microcystin-LR, a toxin from cyanobacteria. Arch. Hydrobiol. 134:417–424.
Erhard, M., von Döhren, H., and Jungblut, P. R. 1999. Rapid identification of the new anabaenopeptin G from Planktothrix agardhii HUB 011 using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Rapid Commun. Mass Spec. 13:337–343.
Erlanger, B. F., Kokowsky, N., and Cohen, W. 1961. The preparation of two new chromogenic substrates of trypsin. Arch. Biochem. Biophys. 95:271–278.
Fastner, J., Erhard, M., and von Döhren, H. 2001. Determination of oligopeptide diversity within a natural population of Microcystis spp. (Cyanobacteria) by typing single colonies by matrix-assisted laser desorption ionization-time of flight mass spectrometry. Appl. Environ. Microbiol. 67:5069–5076.
Fastner, J., Flieger, I., and Neumann, U. 1998. Optimised extraction of microcystins from field samples—A comparison of different solvents and procedures. Water Res. 32:3177–3181.
Fujii, K., Sivonen, K., Naganawa, E., and Harada, K. 2000. Non-toxic peptides from toxic cyanobacteria, Oscillatoria agardhii. Tetrahedron 56:725–733.
Gimenez, A. V. F., Garcia-Carreno, F. L., del Toro, M. A. N., and Fenucci, J. L. 2001. Digestive proteinases of red shrimp Pleoticus muelleri (Decapoda, Penaeoidea): Partial characterization and relationship with molting. Comp. Biochem. Phys. B 130:331–338.
Haney, J. F., Forsyth, D. J., and James, M. R. 1994. Inhibition of zooplankton filtering rate by dissolved inhibitors produced by naturally-occurring cyanobacteria. Arch. Hydrobiol. 132: 1–13.
Hasler, A. D. 1935. The physiology of digestion of plankton crustacea. Biol. Bull. Woods Hole 68:207–214.
Ishitsuka, M. O., Kusumi, T., Kakisawa, H., Kaya, K., and Watanabe, M. M. 1990. Microviridin—A novel tricyclic depsipeptide from the toxic cyanobacterium Microcystis viridis. J. Am. Chem. Soc. 112:8180–8182.
Jakobi, C., Rinehart, K. L., Neuber, R., Mez, K., and Weckessser, J. 1996. Cyanopeptolin SS, a disulfated depsipeptide from a water bloom in Leipzig (Germany): Structural elucidation and biological activities. Phycologia 35:111–116.
Jochimsen, E. M., Carmichael, W. W., An, J. S., Cardo, D. M., Cooksen, S. T., Holmes, C. E. M., Antunes, M. B. D., de Melo, D. A., Lyra, T. M., Barreto, V. S. T., Azevedo, S. M. F. O., and Jarvis, W. R. 1998. Liver failure and death after exposure to microcystins at a hemodialysis center in Brazil. N. Engl. J. Med. 338:873–878.
John, B. K., Plant, D., and Hurd, R. E. 1992. Improved proton-detected heteronuclear correlation using gradient-enhanced z and zz filters. J. Magn. Reson. 101:113–117.
Jongsma, M. A. and Bolter, C. 1997. The adaptation of insects to plant protease inhibitors. J. Insect Physiol. 43:885–895.
Jungmann, D. 1992. Toxic compounds isolated from PCC7806 that are more active to Daphnia than two microcystins. Limnol. Oceanogr. 37:1777–1793.
Kaebernick, M., Rohrlack, T., Christoffersen, K., and Neilan, B. A. 2001. A spontaneous mutant of microcystin biosynthesis: Genetic characterization and effect on Daphnia. Environ. Microbiol. 3:669–679.
Kay, L., Keifer, P., and Saarinen, T. 1992. Pure absorption gradient enhanced heteronuclear single quantum correlation spectroscopy with improved sensitivity. J. Am. Chem. Soc. 114:10663–10665.
Klüttgen, B., Dulmer, U., Engels, M., and Ratte, H. T. 1994. ADaM, an artificial fresh-water for the culture of zooplankton. Freshw. Biol. 28:743–746.
Koppitz, H., Kühl, H., and Kohl, J.-G. 2000. Differences in morphology and C/N-balance between clones of Phragmites australis within a plantation at a degraded fen. Folia Geobot. 35:389–402.
Kotai, J. 1972. Instructions for the Preparation of Modified Nutrient Solution Z8 for Algae. Publication B-11/69. Norsk institutt for vannforskning, Oslo, 5 pp.
Laskowski, M. and Qasim, M. A. 2000. What can the structures of enzyme-inhibitor complexes tell us about the structures of enzyme substrate complexes? BBA-Protein Struct. M. 1477:324–337.
Murakami, M., Sun, Q., Ishida, K., Matsuda, H., Okino, T., and Yamaguchi, K. 1997. Microviridins, elastase inhibitors from the cyanobacterium Nostoc minutum (NIES-26). Phytochemistry 45:1197–1202.
Nagel, W., Willig, F., Peschke, W., and Schmidt, F. H. 1965. über die Bestimmung von Trypsin und Chymotrypsin mit Aminosäure-p-nitroaniliden. Hoppe-Seyler's Z. Physiol. Chem. 340:1–10.
Okino, T., Matsuda, H., Murakami, M., and Yamaguchi, K. 1995. New microviridins, elastase inhibitors from the cyanobacterium Microcystis aeruginosa. Tetrahedron 51:10679–10686.
Piantini, U., Sorensen, O. W., and Ernst, R. R. 1982. Multiple quantum filters for elucidating NMR coupling networks. J. Am. Chem. Soc. 104: 6800–6801.
Pouria, S., de Andrade, A., Barbosa, J., Cavalcanti, R. L., Barreto, V. T. S., Ward, C. J., Preiser, W., Poon, G. K., Neild, G. H., and Codd, G. A. 1998. Fatal microcystin intoxication in a haemodialysis unit in Caruaru, Brazil. Lancet 352:21–26.
Radau, G. 2000. Serine proteases inhibiting cyanopeptides. Pharmazie 55:555–560.
Reinikainen, M. 1997. Acute and Sublethal Effects of Cyanobacteria with Different Toxic Properties on Cladocerean Zooplankton. PhD Thesis. åbo Akademi University, åbo, Finland.
Riemann, B. and Christoffersen, K. 1993. Microbial trophodynamics in temperate lakes. Mar. Microb. Food Webs 7:69–100.
Rohrlack, T., Dittmann, E., Börner, T., and Christoffersen, K. 2001. Effects of cell-bound microcystins on survival and feeding of Daphnia spp. Appl. Environ. Microbiol. 67:3523–3529.
Rohrlack, T., Dittmann, E., Henning, M., Börner, T., and Kohl, J.-G. 1999. Role of microcystins in poisoning and food ingestion inhibition of Daphnia galeata caused by the cyanobacterium Microcystis aeruginosa. Appl. Environ. Microbiol. 65:737–739.
Shin, H. J., Murakami, M., Matsuda, H., and Yamaguchi, K. 1996. Microviridins D-F, serine protease inhibitors from the cyanobacterium Oscillatoria agardhii (NIES-204). Tetrahedron 52:8159–8168.
States, D. J., Haberkorn, R. A., and Reuben, D. J. 1982. A two-dimensional nuclear Overhauser experiment with pure absorption phase in four quadrants. J. Magn. Reson. 48:286–292.
Weckesser, J., Martin, C., and Jakobi, C. 1996. Cyanopeptolins, depsipeptides from cyanobacteria. Syst. Appl. Microbiol. 19:133–138.
Zor, T. and Seliger, Z. 1996. Linearization of the Bradford protein assay increases its sensitivity—Theoretical and experimental studies. Anal. Biochem. 236:302–308.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Rohrlack, T., Christoffersen, K., Hansen, P.E. et al. Isolation, Characterization, and Quantitative Analysis of Microviridin J, a New Microcystis Metabolite Toxic to Daphnia . J Chem Ecol 29, 1757–1770 (2003). https://doi.org/10.1023/A:1024889925732
Issue Date:
DOI: https://doi.org/10.1023/A:1024889925732