The phytases. II. Properties of phytase fractions F 1 and F 2 from wheat bran and the myo-inositol phosphates produced by fraction F 2

Abstract

Wheat bran phytase (EC 3.1.3.8) was fractionated on DEAE-cellulose to yield two fractions, F 1 and F 2. The pH optima (5.6 and 7.2), K m values at pH 5 (2.2·10 −5M and 2·10 −4 M), apparent molecular sizes (both 47 000) and electrophoretic mobilities relative to cytochrome c in starch gels at pH 3.1 (0.47 and 0.58) were determined for F 1 and F 2, respectively. Fraction F 1 shows competitive inhibition by inorganic phosphate ( K i 3·10 −4 M) and F 2 exhibits no detectable inhibition by 10 −2 M phosphate. Myo-inositol phosphates produced in the stepwise dephosphorylation of phytase F 2 have been isolated and characterised as follows: myo-inositol 2-dihydrogen phosphate; d- and l -myo- inositol 1,2-dikis (dihydrogen phosphate); myo-inositol 1,2,3-trikis (dihydrogen phosphate); l -myo- inositol 1,2,3,4-tetrakis (dihydrogen phosphate); myo-inositol 1,2,3,4,6-pentakis (dihydrogen phosphate); l -myo- inositol 1,2,3,4,5-pentakis (dihydrogen phosphate); myo-inositol 1,3,4,5,6-pentakis (dihydrogen phosphate). The results suggest that a variety of phytases are present in biological systems. The initial point of dephosphorylation of phytic acid may occur at the l-i, d-4, 2 or 5 positions of the myo-inositol ring.