Chymosin is the key enzyme that can specifically cleave κ-casein and manufacture cheese.In this study,we expressed the bovine prochymosin in Escherichia coli,purified and activated the recombinant prochymosin,measured its milk-clotting activity,then analyzed the enzymatic properties of chymosin.The results showed that the prokaryotic expressed prochymosin accounted for about 66.3% of the total bacterial protein,the purified prochymosin was about 200 mg / L of culture liquid,and chymosin activity was 600 000 SU / g after prochymosin activation.The optimal temperature range of milk-clotting activity of recombinant chymosin was 57 ~ 62 ℃,and the chymosin was relatively stable in pH 2 ~ 7 and below 40℃.Metal ions such as Al3 +,Fe3 +and Cu2 +significantly increased the activity of chymosin,however,pepsin inhibitor pepstatin A had an obvious inhibitory effect on the activity of chymosin.It would provide a high expression strain for the industrial production of chymosin.