Abstract: In order to improve the cell concentrations and L- aspartase activity durting the L- asparate production of Escherichia coli HY- 05 C,the culture medium and culture conditions were optimized. First of all,the orthogonal experiment was employed to optimize the medium compositions and the optimal fermentation medium was obtained( g / L) : ammonium fumarate 10,corn steep liquor powder 8,yeast powder 2,peptone 7,Na Cl 5,KH₂PO₄1,Mg SO₄ 0.2.Furthermore,the initial p H and inoculum size which had significant influence on bacteria concentration and enzyme activity were investigated,and the optimal p H and inoculum size were determined as 6.0 and 1‰,respectively.Finally,the enzymatic conversion process of E.coli HY-05 C was confirmed using the optimal medium and the culture conditions. Compared with the initial cultual conditons,its conversion efficiency was improved by150%. Also,it verified the feasibility that the increasing cell concentrations via the optimization of fermentation conditions could enhance the conversion efficiency of L- asparatase in E.coli HY-05 C.