Mechanism-based irreversible inhibitors of cytokinin dehydrogenase
Jeffrey C Suttle1, Rene Mornet
1US Department of Agriculture, Agriculture Research Service, Northern Crop Science Laboratory, Fargo, North Dakota 58105-5677, USA. suttlej@fargo.ars.usda.gov
Abstract:
The effects of three N(6)-substituted aminopurine derivatives containing either allenic or acetylenic side-chains on in vitro and in vivo cytokinin dehydrogenase (CKX; EC 1.5.99.12) activities were determined. At concentrations < or = 100 microM, the acetylenic derivative (HA-2) had no effect on in vitro CKX activity. In contrast, the two allenic derivatives (HA-1, HA-8) inhibited in vitro CKX activity in a dose-dependent manner with 50% inhibition occurring at HA-1 and HA-8 concentrations of 9.0 and 0.4 microM (respectively). HA-8 inhibited the degradation of both the free bases and ribosides of N6-(2-isopentenyl)adenine and zeatin. Pretreatment with HA-8 inhibited CKX activity in both a time- and concentration-dependent manner. In contrast to the reversible phenylurea inhibitor N-(chloro-4-pyridyl)-N'-phenylurea, inhibition of CKX activity by HA-8 was not relieved by 24 h of dialysis. Both HA-1 and HA-8 (but not HA-2) inhibited the metabolism of exogenous [3H]-N(6)-(2-isopentenyl)adenosine in excised aseptic potato (Solanum tuberosum) leaves. These results demonstrate that HA-8 is a mechanism-based irreversible (suicide) inhibitor of CKX and indicate that it may be useful in determining the role of CKX in cytokinin homeostasis in planta.
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