Colorimetric protein phosphatase inhibition assay of laboratory strains and natural blooms of cyanobacteria: comparisons with high-performance liquid chromatographic analysis for microcystins.

Microcystins are cyclic heptapeptide hepatotoxins commonly produced by bloom-forming genera of cyanobacteria. These toxins are potent and specific inhibitors of protein phosphatases 1 and 2A. We have optimised a rapid, simple and sensitive colorimetric protein phosphatase 1 inhibition assay, utilising the activity of protein phosphatase 1 as expressed in a recombinant strain of Escherichia coli, towards the chromogenic substrate, p-nitrophenyl phosphate. A standard curve for the inhibition of protein phosphatase 1 by microcystin-LR was constructed with an IC50 of about 38 ng ml-1 and a limit of detection of 10-20 ng ml-1. Twenty-three laboratory-grown strains and 25 natural bloom samples of cyanobacteria were analysed by high-performance liquid chromatography for microcystins and by the protein phosphatase 1 inhibition assay. Agreement for the microcystin contents of the samples detected by high-performance liquid chromatography and the protein phosphatase 1 inhibition assay showed good correlation (R2 > 0.93, P < 0.0001). The suitability of the colorimetric protein phosphatase 1 inhibition assay as a screen for cyanobacterial microcystins is discussed.

[1]  R. Harrison,et al.  Detection analysis and risk assessment of cyanobacterial toxins , 1996 .

[2]  L. Lawton,et al.  Isolation and characterization of microcystins from laboratory cultures and environmental samples of Microcystis aeruginosa and from an associated animal toxicosis. , 1995, Natural toxins.

[3]  W. Carmichael,et al.  Use of a colorimetric protein phosphatase inhibition assay and enzyme linked immunosorbent assay for the study of microcystins and nodularins. , 1994, Toxicon : official journal of the International Society on Toxinology.

[4]  G. Codd,et al.  Cyanobacterial toxins and human health , 1994 .

[5]  G. Codd,et al.  Toxic blooms of cyanobacteria in Lake Alexandrina, South Australia — Learning from history , 1994 .

[6]  A. Sim,et al.  Protein phosphatase activity in cyanobacteria: consequences for microcystin toxicity analysis. , 1993, Toxicon : official journal of the International Society on Toxinology.

[7]  M. Browner,et al.  Expression of the catalytic subunit of phosphorylase phosphatase (protein phosphatase-1) in Escherichia coli. , 1992, The Journal of biological chemistry.

[8]  J. Zwiller,et al.  Characterization of microcystin-LR, a potent inhibitor of type 1 and type 2A protein phosphatases. , 1990, The Journal of biological chemistry.

[9]  Philip R. Cohen,et al.  Cyanobacterial microcystin‐LR is a potent and specific inhibitor of protein phosphatases 1 and 2A from both mammals and higher plants , 1990, FEBS letters.

[10]  G. Codd,et al.  Cyanobacterial Toxins in Water , 1989 .

[11]  Dudley H. Williams,et al.  The structure of cyanoginosin-LA, a cyclic heptapeptide toxin from the cyanobacterium Microcystis aeruginosa , 1985 .

[12]  I. Falconer,et al.  Evidence of liver damage by toxin from a bloom of the blue‐green alga, Microcystis aeruginosa , 1983, The Medical journal of Australia.

[13]  G. Cohen-bazire,et al.  Purification and properties of unicellular blue-green algae (order Chroococcales). , 1971, Bacteriological reviews.