Objective: The cell disruption step in recombinant protein production is critical for the quantity and quality of the protein obtained. In this study, the effectiveness of sonication and high-pressure methods was tested on E. coli BL21 cells producing recombinant SpdAZ protein.

Methods: This study assessed the effectiveness of sonication and high-pressure techniques on E. coli BL21 cells expressing recombinant SpdAZ protein.Cell disruption was performed using high pressure (5000, 10000, and 20000 psi) and sonication (50 kHz) methods, with 3, 6, and 9 repetitions. A control group with no treatment was included to evaluate the disruption efficiency. The degree of cell lysis was measured by counting live cells after serial dilution and plating, and enzyme activity was tested with DNase agar. DNA fragmentation and protein concentration were also analyzed.

Results: Both methods showed a decrease in live cell counts as the number of repetitions increased. After 9 cycles, sonication reduced live bacteria to 1016 cfu/ml, while high pressure (20000 psi) reduced it to 108 cfu/ml. Enzyme activity, as indicated by the DNase test, revealed a 23 mm zone for high-pressure lysates compared to 7 mm for sonication. DNA fragmentation analysis showed that high-pressure treatment was 19% more effective. Protein concentrations reached 16570 mg/ml for the high-pressure method versus 12320 mg/ml for sonication.

Conclusion: The disruption of cells is crucial for maintaining enzyme activity and protein structure in recombinant protein production. In the cell disruption phase, the high-pressure method was found to be superior to sonication in terms of live cell count, enzyme activity, and total protein yield.