| Budget Amount *help |
¥3,400,000 (Direct Cost: ¥3,400,000)
Fiscal Year 2003: ¥700,000 (Direct Cost: ¥700,000)
Fiscal Year 2002: ¥2,700,000 (Direct Cost: ¥2,700,000)
|
|---|
| 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.
|
