title: Inhibition equivalency factors for microcystin variants in recombinant and wild-type protein phosphatase 1 and 2A assays.
abstract: In this work, protein phosphatase inhibition assays (PPIAs) have been used to evaluate the performance of recombinant PP1 and recombinant and wild-type PP2As. The enzymes have been compared using microcystins-LR (MC-LR) as a model cyanotoxin. Whereas PP2ARec provides a limit of detection (LOD) of 3.1 μg/L, PP1Rec and PP2AWild provide LODs of 0.6 and 0.5 μg/L, respectively, lower than the guideline value proposed by the World Health Organization (1 μg/L). The inhibitory potencies of seven MC variants (-LR, -RR, -dmLR, -YR, -LY, -LW and -LF) have been evaluated, resulting on 50 % inhibition coefficient (IC50) values ranging from 1.4 to 359.3 μg/L depending on the MC variant and the PP. The PPIAs have been applied to the determination of MC equivalent contents in a natural cyanobacterial bloom and an artificially contaminated sample, with multi-MC profiles. The inhibition equivalency factors (IEFs) have been applied to the individual MC quantifications determined by liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis, and the estimated MC-LR equivalent content has been compared to PPIA results. PPIAs have demonstrated to be applicable as MC screening tools for environmental applications and to protect human and animal health.
authors.affiliation: IRTA, Carretera de Poble Nou km 5.5, 43540, Sant Carles de la Ràpita, Spain.
authors.forename: Diana
authors.initials: D
authors.lastname: Garibo
authors.forename: Cintia
authors.initials: C
authors.lastname: Flores
authors.forename: Xavier
authors.initials: X
authors.lastname: Cetó
authors.forename: Beatriz
authors.initials: B
authors.lastname: Prieto-Simón
authors.forename: Manel
authors.initials: M
authors.lastname: Del Valle
authors.forename: Josep
authors.initials: J
authors.lastname: Caixach
authors.forename: Jorge
authors.initials: J
authors.lastname: Diogène
authors.forename: Mònica
authors.initials: M
authors.lastname: Campàs
mesh_terms: D002853:Chromatography, Liquid; D000458:Cyanobacteria / Q000737:chemistry; D004791:Enzyme Inhibitors / Q000737:chemistry*; D006801:Humans; D013058:Mass Spectrometry; D052998:Microcystins / Q000737:chemistry*; D054645:Protein Phosphatase 1 / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D054648:Protein Phosphatase 2 / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D004791:Enzyme Inhibitors; D052998:Microcystins; C078588:microcystin; D054645:Protein Phosphatase 1; D054648:Protein Phosphatase 2