Molecular identification and growth optimization of protease-producingSerratia isolated from soil

Background and Objective: Proteases are the most important enzymes used in industry, laboratory, andmedical purposes. A large number of researches show that fungi and bacteria are the main sources ofprotease production. Therefore, considering the importance of finding enzyme-producing microorganismsfrom new natural sources, the purpose of this study is to isolate and identify the protease-producingbacteria isolated from the soil of different regions of Iran.Methods: In the present study, after collecting soil from different regions of Hamadan, Fars,Chaharmahal and Bakhtiari provinces, preliminary screening was done among ۲۰ isolated bacterialisolates. First, the obtained samples were cultured on skimmed milk agar medium in order to evaluate theproduction of protease. Then, they evaluated the diameter of the transparent halo created around thebacterial growth area, and the isolates producing the largest halo were selected for identification and amore detailed investigation. In the next step, using casein substrate and UV absorption in the range of ۲۸۰nm, protease enzyme activity was measured, and then identification is done through a moleculartechnique based on ۱۶s rDNA detection. Finally, Optimizing conditions for growth such as severalcompounds added to the culture medium, pH, and different temperature ranges of the selected samplewere tested. Also, to determine the maximum growth, a spectrophotometer was used at a wavelength of۶۰۰ nm.Results: In this research, among ۲۰ bacterial isolates, one isolate with the highest protease activity wasselected for further investigations. The results of sequencing and aligning in the NCBI database showed ahigh similarity of ۹۸% to the genus Serratia. Also, the results showed that it would grow by using acombination of ۳ grams of yeast extract, ۵ grams of peptone, and ۱۰ CC of sugar beet molasses per literin the culture medium at the maximum level. In addition, it showed the highest protease activity in thetemperature range of ۲۸ to ۳۲ degrees Celsius. Enzyme activity was the highest in the pH range of ۵ to ۸without no significant difference.Conclusion: In agreement with our findings there are many reports about the potential of using proteasesof Serratia. The main advantage of using this protease is its activity in a relatively wide range of pH,which facilitates is its use. Another features of this protease its relatively simple culture medium.

Keywords:

Molecular identification , Optimization , Protease , Serratia