Water stress inhibits p-nitrophenyl phosphate hydrolysis activity of the plasma membrane H+-ATPase from soybean hypocotyls

Abstract

The influence of water stress on ATP and p-nitrophenyl phosphate (PNPP) hydrolysis by plasma mem-brane ATPases was investigated using plasma membrane vesicles purified from soybean hypocotyls by the sucrose gradient centrifugation method. Results showed that ATPase activity was reduced after 10% polyethylene glycol (PEG) 6000 treatment for 12 h. Water stress also moved the optimal pH from 6.5 to 7.0. A significant decrease in PNPP hydrolysis was observed under PEG treatment. The Km for PNPP hydrolysis was shifted from 2.3042 0.0009 to 2.5048 0.0346 mmol L –1 . Moreover, PNPP hydrolysis was more sensitive to vanadate after PEG treatment, while inhibition of ATP hydrolysis by hydroxylamine was not affected. Our experimental results indicated that water stress changed the catalytic mechanism of the plasma membrane H + -ATPase through affecting the dephosphorylation process catalysed by its phosphatase domain.