Homa Noura - Iranian Research Organization for Science and Technology
Nahid Bakhtiari - Iranian Research organization for science and technology
ali Sheykhinejad - Iranian Research Organization for Science and Technology

Background and Aim: Bacterial expression systems for heterologous protein production are attractive because of their ability to grow rapidly and at high density on inexpensive substrates. However, the rational choice of the adequate promoter system and host for a specific protein of interest remains difficult. A given gene product whether foreign or native can be toxic to the cell when expressed in large quantities. In this project, we observed the effect of two antibacterial recombinant proteins, on E. coli Bl۲۱ (DE۳) growing process in the auto-inducing medium. One of the recombinant proteins was phage native lysin and other one was engineered type of this enzyme. Lysins are phage enzymes that can hydrolase peptidoglycan layer of bacterial membrane.Methods: An auto-inducing medium have lactose as inducer sugar was used for preventing of basal expression and increasing the accuracy of investigation about the effect of external protein expression on bacterial host growth. For pre-culture, a non-inducing media have no inducer sugar and made entirely from purified ingredients was used to minimize contamination probability by inducing agents. Auto-inducing cultures with ۵۰ µg/ml kanamycin were inoculated with one- thousandth volume of saturated pre-culture medium. Bacterial cultures were grown over ۲۴ hours at ۳۷ºC in a shaker incubator with shaking rate of ۲۲۰ rpm. Bacterial growth was monitored by measuring the optical density at ۶۰۰ nm (OD۶۰۰nm) per hour. Growth curves were plotted for all bacteria have heterologous recombinant protein.Results: The results showed that the presence of heterologous recombinant engineered lysin was prevented the bacterial growth significantly and just continued until an OD۶۰۰ ~۶ comparing with OD۶۰۰ ~۱۶ for native lysin.Conclusion: The engineered lysin could destroy E.coli from inside of the cell. Therefore, the antibacterial effect of an engineered enzyme is absolutely noticeable in comparison to intact native protein also in this stage.

antibacterial, lysin, E.coli, growth