Is Industrial Crystallization a Suitable Process for Protein Purification? – A Survey
Advancing the chemical engineering fundamentals
Keynote Lectures: Theme-2
Keywords: protein purification, crystallization, enzymatic activity, downstream processing
With the growing importance of protein products, demand for highly selective and cost effective separation methods with low environmental impact will increase. Industrial crystallization has the potential to satisfy all three requirements. However, for protein products, crystallization in the form of precipitation by salting-out is rarely used as anything other than an initial fractionating step prior to further purification, usually by chromatography. In contrast, protein crystal growth is well established for generating high-quality single crystals for structural studies. Despite being well established, the technique nonetheless relies upon empirical screening approaches when a new protein is to be crystallized. In part this is due to the absence of a general understanding of the factors influencing protein crystallization, which in turn is a consequence of the compexity of the problem: in addition to the "classic" crystallization parameters (temperature, pressure, concentration), account has to be taken of the effect of the precipitating agent required (its nature and concentration) as well as of the pH at which crystallization is carried out.
In the case of industrial crystallization, there is the added complication of the raw materials from which the protein product is to be isolated, either renewable resources or fermentation broths. Either of these represent complex systems containing innumerable different compounds which may influence the processing requirements of the target protein.
This contribution will provide an overview of the state-of-the-art in protein crystallization with a view to developing large-scale processes for purification. Case studies based upon research carried out in our group will illuminate the difficulties that can be expected and which have to be addressed in the development of a process. Particular attention will be paid to mixing of the protein solution with the precipitant and to characteristics of enzymes, in particular their catalytic activity. The activity is an important characteristic and it is desirable to maintain the catalytic power of an enzyme during purification. In addition, the activity can be employed as a measure of the quality of a purification process. The enzyme urease, isolated from the jack bean, will be used as an example to highlight the effect of process conditions upon the activity. In addition, alternatives to standard batch crystallization by salting-out will be discussed.
Presented Wednesday 19, 17:05 to 17:45, in session Keynote Lectures: Theme-2 (T2-K4).
