To obtain a high level expression of phytase with favorable characteristics, a codon-optimized phytase gene from Citrobacter freundii was synthesized and transferred into Pichia pastoris. Small-scale expression experiments and activity assays were used to screen positive colonies. After purified by Ni2+-NTA agarose affinity column, the characterizations of the recombinant phytase were determined. The recombinant phytase (r-phyC) had two distinct pH optima at 2.5 and 4.5 and an optimal temperature at 50 degrees C. It retained more than 80% activity after being incubated under various buffer (pH 1.5-8.0) at 37 degrees C for 1 h. The specific activity, K-m, and V-max values of r-phyC for sodium phytate were 2,072 +/- 18 U mg(-1), 0.52 +/- 0.04 mM, and 2,380 +/- 84 U mg(-1) min(-1), respectively. The enzyme activity was significantly improved by I mM of K+, Ca2+, and Mg2+. These characteristics contribute to its potential application in teed industry.