Plant protein tyrosine phosphatases (PTPs) are important enzymes in regulating cellular responses to redox change through their reversible inactivation under oxidative conditions. In contrast to previous results showing that soybean seeds acid phosphatases were not significantly affected by hydrogen peroxide, pervanadate, p-chloromercuribenzoate and Cu²⁺, recent reports have suggested that soybean PTPs were relatively sensitive to the presence of oxidizing compounds. The aim of this work was to demonstrate that preincubation effects are responsible for the oxidative inactivation of soybean seeds acid phosphatases.
The enzyme activity was determined by measuring the p-nitrophenol released in the presence of p-nitrophenylphosphate as substrate. The acid phosphatase activity was measured after 10 minutes of enzyme preincubation with the oxidizing compounds. The enzyme activity decreased 72%, 68% and 95% in the presence of Cu²⁺, hydrogen peroxide and pervanadate, respectively. An enzyme preincubation for 10 minutes, in the presence of GSH and before treatment with Cu²⁺, showed about two times more activity than without GSH, suggesting an enzyme protection promoted by GSH, against the oxidation. Our results confirmed that, like other PTPs, the enzyme from soybean seeds had also one or more essential catalytic cysteine residues. The necessity of enzyme preincubation with the inhibitors could be explained by the relative inaccessibility to the modifying agents, for instance, due to the presence of carbohydrates. It is worthwhile mentioning that oxidative inhibitions of castor bean seeds acid phosphatase, that also contained carbohydrate moiety, was not dependent on preincubation. The presence of cysteine residues at PTP active site is important for the redox regulation of this class of enzymes.
Financial Support: CAPES, CNPq, FAPESP and FAEPEX/SAE/UNICAMP.