根据密码子的偏好性,采用PTDS方法化学合成了一个来源于Yersinia kristensenii的新型植酸酶基因(YkAPPAS),并且作为外分泌蛋白在毕赤酵母GS115中成功表达.重组植酸酶YkAPPAS的蛋白分泌量可达到106.73μg/mL.对重组植酸酶的酶学性质研究表明:该酶的比活力为2 330 U/mg;最适pH为4.5和6.0;最适温度为45℃;100℃处理5 min后其剩余活性为47.55％;90℃处理5 min后其剩余活性为56.75％;在pH3-12时的活性均在30％以上.Li+、EDTA对酶活有一定的激活作用;Zn2、Cu2对酶活有一定的抑制作用.重组植酸酶还具有良好的抗胃蛋白酶和胰蛋白酶水解的特性.%A novel phytase gene YkAPPAS from Yersinia kristensenii was chemically synthesized by the PCR-based two-step DNA synthesis according to yeast codon preference and successfully expressed as secreted protein in Pichia pastoris GS115.The recombinant YkAPPAS protein secretion yield could reach 106.73 μg/mL.The research of its enzymological property showed that its specific activity was 2 330 U/mg;Its optimal pH was at 4.5 and 6.0 and the optimal temperature was 45 ℃;Its residual activity was 47.55％ after 5-min treatment at 100 ℃ and 56.75％ after 5-min treatment at 90 ℃;Between pH 3-12 its activity was above 30％.To some extent its enzyme activity could be activated by Li + and EDTA but inhibited by Zn2+ and Cu2+.It had a good ability to resist hydrolyzation by pepsin and trypsin.
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