Abstract: Objective:To determine whether the activity of subtilisin QK produced by Bacillus subtilis from different strains is related to the amino acid site mutation encoding its sequence. Methods:The total DNA of 15 strains was used as a template. The 16S rRNA of bacteria was used as an universal primer for PCR amplification. The products were sequenced and input into NCBI to determine that they were all B. subtilis. The QK genes of 15 strains were amplified and sequenced. The protein sequences corresponding to the submitted QK gene in GenBank were aligned to find different amino acid sites. Fibrin plate method was used to determine the activity of the subtilisin produced by these strains. BCA protein assay kit was used to determine the total protein content of the fermentation broth. The unit enzyme activity was calculated. Results:According to the mutation sites, 15 strains were divided into four groups. The mutation site of group A was S184T. The unit enzyme activity of group A was the highest (p<0.05). The mutation sites of group B were Q90K, N193S, S236T, N365S, and the mutation sites of group C were N193S, S236T, A289V. The results showed that there was no significant difference between these two groups (p>0.05).But the enzyme activity in groups B and C was significantly lower than that of group A (p<0.05). The mutation sites of group D were Q90K, N193S, S236T, A289V, and the unit enzyme activity was significantly lower compared with that of group A, B and C (p<0.05).Conclusion:There was an obvious relationship between the unit enzyme activity of subtilisin QK and amino acid mutation sites. Among these amino acid mutation sites, the sites of S184T and N365S might promote the activity of unit enzyme. And the site A289V might have an inhibitory effect. These sites might be located at the active center of subtilisin QK.