Viscosity Reduction through Co-Expression of Nuclease for Improved Clarification

Alternative strategies are required in downstream processing of biopharmaceuticals to reduce the cost of manufacturing. The use of host cell engineering to achieve step-change improvement in the efficiency of unit operations involved in downstream processing is demonstrated in this paper. Nucleic acids co-released during product recovery contribute to the increased viscosity of the process stream. High viscosity of the product stream decreases the efficiency of the unit operations involved such as centrifugation, filtration, and chromatography. Staphylococcal aureus nuclease directed to the periplasm was co-expressed in E. coli to reduce the viscosity of process stream through auto-hydrolysis of nucleic acids. The reduction in the viscosity and the resultant step-change improvements in clarification performance were quantified.

Viscosity reduction of up to 75 % was achieved while avoiding micronisation of cell debris at the same time. The viscosity reductions were comparable and even slightly better than commercially available nuclease. The autohydrolysis of DNA enabled preparation of homogenates at increased biomass concentration resulting in concentrated product stream. The less viscous process stream dramatically altered the clarification performance in scale-down mimics of continuous disc-stack centrifugation. Laboratory scale data indicated that a 4 fold reduction in settling area of disc stack centrifuge can be expected due to less viscous product stream. The details of the step-change improvements achieved will be discussed in the paper.