Bacterial cell disruption: a crucial step in protein production.

Recombinant protein production significantly improved in the past three decades. Novel expression systems were developed, growth conditions optimised and the technology and thus monitoring and analysis significantly enhanced. However, the studies of bacterial cell disruption were more or less neglected. The existing methods were acceptable until the final product of protein production was soluble and pure protein. However recently, inclusion bodies (IBs) as whole protein particles were also recognised as the final product. Classical methods for bacterial cell disruption are therefore not always suitable, sufficient or even appropriate for isolation of such particulate material. Some of the currently existing methods for bacterial cell disruption were recognised as damaging for the structure of IBs, while sonication was even found harmful for the recombinant protein. The powers needed for disruption of the bacterial cells damage the recombinant proteins and thus their biological activity significantly reduces. Furthermore, the classical isolation methods enable disruption of majority of the bacterial cells and this is enough for isolation of soluble proteins, yet it is not adequate for isolation of particulate material. While remaining bacterial cells sediment together with the IBs, they represent impurity. The need for isolation of cell-free IBs was therefore revealed in the recent studies, because only pure IBs can be used as nanoparticles in further biomedical applications. Therefore it is time to consider, redesign, optimise or even develop new alternative methods that would enable isolation of pure, structurally intact and biologically active particles. Two such alternative methods that enable isolation of bacterial free, active protein particles were developed recently.