Purification of Proteins and the Disruption of Microbial Cells

TLDR

Intracellular proteins are increasingly valuable for enzyme technology and recombinant production, making their release from microorganisms—a first step in isolation—critical, and large‑scale disruption is typically achieved with mechanical disintegrators originally designed for other industries. The study aims to enhance the design and performance of these disintegrators for cell disintegration, targeting uniform cell exposure, minimal protein shear, high yields, and avoidance of overly small cell‑wall fragments. Both high‑speed bead mills and high‑pressure homogenizers were evaluated across various microbial cells, with operating parameters examined for their effects on protein solubilization and enzyme yield.

Abstract

Abstract Intracellular proteins with catalytic or biological activity are of growing importance for developments in enzyme technology, as well as for the production of mammalian proteins by recombinant‐DNA technology. The release of these proteins from microorganisms is an important unit operation, as it is the first step in their isolation. Gram‐scale disruption of microorganisms can be performed by a variety of established methods based on chemical, enzymatic, physical, or mechanical principles. For the large scale disruption of microorganisms, mechanical disintegrators, such as high‐speed agitator bead mills or high‐pressure industrial homogenizers, are commonly employed. Both types of equipment were designed originally for other tasks; in the paint industry or in the milk industry, respectively. Therefore, it appeared necessary and possible to improve design and performance for the application in cell disintegration. The goal is a uniform exposure of the microbial cells and a minimal exposure of solubilized protein to high shear forces, in order to obtain high yields and to avoid the generation of too small cell wall fragments, which are difficult to separate. Both types of machines have been investigated for the disintegration of different microbial cells and the influence of the operating parameters analyzed on protein solubilization and enzyme yield. We will summarize the state of the art and discuss new data to illustrate trends in process development.