FACTORS DETERMINING MORE EFFICIENT LARGE-SCALE RELEASE OF A PERIPLASMIC ENZYME FROM ESCHERICHIA-COLI USING LYSOZYME

Large scale use of lysozyme for periplasmic release has been impeded b y the cost of the pure enzyme and its subsequent presence as a contami nant in later downstream processing steps. In this paper, we discuss t he use of lysozyme for pilot scale recovery of a periplasmic enzyme fr om E. coli. The effects of concentration of sucrose, lysozyme and cell s on periplasmic enzyme release were examined. Lysozyme concentration can be reduced 5-fold from previous reports and a reduction in sucrose concentration from 20 to 15% (w/v) allows an improvement in centrifug al harvesting by reducing viscosity. High levels of release were still achieved using this technique and further improvements in yield were obtained by optimising other components of the releasing mixture. Resu lts show that some release is still achieved in circumstances where no lysozyme use is possible. Results also indicate that a substantial pr oportion (up to 70%) of lysozyme remains bound to the cellular debris after its action and is removed with this material. (C) 1997 Elsevier Science B.V.