Glycolytic intermediates as substrates of soybean acid phosphatase isoforms

The kinetic properties of four isoforms of acid phosphatase purified from m ature soybean (Glycine max) seeds were studied using glycolytic metabolites as substrates. The isoforms AP1, AP3A and AP3B presented maximal activitie s around pH 4.0, and AP2 at pH 6.0, for phosphoenolpyruvate (PEP) as a subs trate. With glucose-6-P (G6P) and fructose-6-P (F6P) maximal activities wer e observed at pH 5.5 for the AP3A and AP3B isoforms. The acid phosphatases presented the following apparent K-m values, at the corresponding optimum p H values: AP1 (p-nitrophenylphosphate (pNPP): 0.49, PEP: 0.23 mM); AP2 (pNP P: 0.38, PEP: 0.47 mM); AP3A (pNPP: 0.20, PEP: 0.10, G6P: 0.30, F6P: 0.16 m M) and AP3B (pNPP: 0.086, PEP: 0.078, G6P: 0.31, F6P: 0.33 mM). Maximal spe cificity constant (V-max/K-m) was obtained for the isoform:AP1, with PEP as a substrate. Independent of the substrates, the reactions catalyzed by the soybean acid phosphatase isoforms were potently inhibited by molybdate and to a lesser extent by Zn2+. Inhibitions were also observed in the presence of fluoride, with PEP as a substrate, and by Cu2+, and p-chloromercuribenz oate (pCMB) with G6P and F6P as substrates. Our results suggest that the Fo ur acid phosphatase forms could play important roles in plant metabolism, a cting on key glycolytic intermediates. (C) 1999 Elsevier Science Ireland Lt d. All rights reserved.