A number of phytase genes and proteins have been identified from microorganisms and plants after the isolation of the very first phytase (PhyA) protein and DNA sequence from Aspergillus niger. It remains unclear whether phytase is expressed in animal tissues. PhyA and most fungal phytases have molecular mass ranging from 80 120 kDa, with 10 or so N-glycosylation sites in the approximately 1.4-kb DNA sequences. The average molecular masses of most bacterial phytases range from 40 to 55 kDa. Plant phytases isolated from corn, wheat, lupine, oat, or barley have molecular sizes ranging from 47 to 76 kDa. Most identified phytases, but not all, belong to a group of histidine acid phosphatases (HAPs) that feature the conserved active site hepta-peptide motif RHGXRXP and the catalytically active dipeptide HD. This group of phytases catalyzes phytic acid hydrolysis in a two-step mechanism via a nucleophilic attack from the histidine in the active site of the enzyme to the scissile phosphoester bond of phytic acid. In general, fungal phytases (E.C. 126.96.36.199) initiate the splitting of the phosphate group at the C1 or C3 carbon of the inositol ring, and are thus called 3-phytases, whereas plant phytases (E.C. 188.8.131.52) act preferentially at the C6 carbon, and are named 6-phytase. However, phytases isolated from Escherichia coli, Lupinus albus, or Pen-iophora lycii are exceptions to this rule. Interestingly, a soybean phytase is a purple acid phosphatase with a dinuclear iron iron or iron zinc center in the active site. The phytase from Bacillus subtilis has a six-bladed folding scaffold, and calcium ion can affect its thermo-stability and catalysis. As a whole, phytases show a strong ability to cleave equatorial phosphate groups, but a limited ability to hydrolyze axial phosphate groups. The optimum pH for most known phytases is in the range of 4.5 6. Exceptions are phytases from mung bean, Enterobacter sp., or B. subtilis that have their pH optimum in the neutral to alkaline range. The temperature optimum of most plant and microbial phytases ranges from 45 65°C, higher than body temperatures of animals (37 40°C). Phytase activity unit is defined by the amount of inorganic phosphate released per minute from a selected substrate under certain pH and temperature. In the case of A. niger PhyA, the activity is determined in 0.05 0.2 M citrate or acetate buffer, pH 5.5, at 37°C; and one unit equals the amount of enzyme that releases 1 mmol of inorganic phosphorus per minute from sodium phytate. The biochemical properties of the currently available phytases for animal feeding are summarized in Table 1.
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