Gene Cloning, Expression and Characterization of a Novel Phytase from Hafnia alvei

Abstract:

A gene appA encoding a novel phytase was firstly cloned from Hafnia alvei by PCR and sequenced. The gene was consisted of 1335 bp, encoding 444 amino acids. The calculated molecular weight of the mature APPA was about 45.2 kD. The gene appA was expressed in E. coli BL21(DE3). Recombinant APPA was purified and its enzymatic properties were determined. The optimum pH for the enzyme was 4.5 and the optimum temperature was 60℃.The pH stability of r-APPA is good, the relative phytase activity was above 80% after treated in buffers of pH2.0～10.0. The specific activity of r-APPA is 356.7 U/mg, and the K_m value was 0.49 mmol/L and V_max of 238 U/mg. The enzyme showed resistance to pepsin and trypsin treatment.