2003 Fiscal Year Final Research Report Summary
study of Physiological activity of Hepatotoxins, Mirocystins Produced by freshwater Cyanobacteria
| Project/Area Number |
14572115
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Environmental pharmacy
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| Research Institution | Meijo University |
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Principal Investigator |
HARADA Ken-ich Meijo University, Faculty of pharmacy, professor, 薬学部, 教授 (90103267)
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| Co-Investigator(Kenkyū-buntansha) |
KAZUAKI Kawai University of Ouupational and Environmental Heal, associate professor, 医学部, 助教授 (60161262)
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| Project Period (FY) |
2002 – 2003
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| Keywords | Microcystin / Protein phosphatase / Okadaic acid / Oxidative stress / Mass spectrometry / Phosphorylation of proteins / Binding proteins / Hepato toxicity |
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| Research Abstract |
Microcystin-LR (MCLR) produced by freshwater cyanobacteria is potent hepatotoxin, and inhibits protein serine/threonine phosphatases 1 and 2A (PP1 and PP2A). A toxicological mechanism has not been elucidated, so that an appropriate treatment has not been yet established for Microcystin toxicosis. In order to clarify this acute toxicity mechanism by MC we have searched a targeted compound and related compounds for MCLR in the liver during the toxicological process. In this study, we always compared the behavior with MC and that of okadaic acid (OA), which does not show such toxicity but the similar PP inhibitory activities.
1.Mt-binding proteins were comprehensively searched in mouse livers using MC affinity column, two-dimensional gel electrophoresis and MALD1-TOFMS. The results obtained from the competitive inhibition experiments using the affinity chromatography with OA indicated that the complex of PP1 with the inhibitory subunit NIPP1 was only detected as the MC-binding proteins.
2.In order to investigate proteins increased in phosphorylation in mouse livers, resulting phosphoproteins were chemically modified with cleavable biotin reagent on a phosphate moiety, purified by an avidin column, and then analyzed by ESI-LC/MS/MS. Consequently, a phosphoprotein, formyltetrahydrofolate dehydrogenase was only detected in both the MCLR-and OA-treated mice. These results suggested that the phosphorylation was caused by PP2A inhibition, which was not related to the liver injury.
2.In order to investigate proteins increased in phosphorylation in mouse livers, resulting phosphoproteins were chemically modified with cleavable biotin reagent on a phosphate moiety, purified by an avidin column, and then analyzed by ESI-LC/MS/MS. Consequently, a phosphoprotein, formyltetrahydrofolate dehydrogenase was only detected in both the MCLR-and OA-treated mice. These results suggested that the phosphorylation was caused by PP2A inhibition, which was not related to the liver injury.
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