Comparative study of two trehalase activities from Fusarium oxysporum var lini

Acid and neutral trehalase activities (optimum pH of 4.6 and 6.8, respectively) from Fusarium oxysporum var. lini were studied separately through partial isolation by ammonium sulfate precipitation followed by ion-exchange chromatography on DEAE-Sephacel for neutral enzyme, or using some of their differential properties. Acid activity was unaffected by 1 mM of Ca2+, Mg2+, Mn2+, Ba2+, or EDTA. Contrarily, the neutral enzyme was activated by Ca2+ with an apparent K-a of 0.15 mM; was inhibited by EDTA, Zn2+, Hg2+, or Mg2+-ATP; and showed an increase in activity by the raise of buffer ionic strength or by the addition of 100 mM KCI. Acid and neutral enzymes have, respectively, an apparent optimum temperature of 45 and 30 degrees C, an apparent K-m for trehalose of 0.43 and 8.45 mM, and an apparent M(r) of 160 000 and 100 000 (by glycerol gradient ultracentrifugation). Acid trehalase was specifically inhibited by acetate buffer and more stable at 50 degrees C than the neutral enzyme. Neutral enzyme exhibited a pi of 6.2 by isoelectric focusing. Contrary to neutral trehalases from other fungi, the enzyme from Fusarium oxysporum var. lini was not activated in crude extract by treatment with Mg2+-ATP in the presence of cAMP and not inactivated by alkaline phosphatase from Escherichia coli.