A new protein folding screen: Application to the ligand binding domains ofa glutamate and kainate receptor and to lysozyme and carbonic anhydrase

Production of folded and biologically active protein from Escherichia coli derived inclusion bodies can only be accomplished if a scheme exists for in vitro naturation. Motivated by the need for a rapid and statistically mean ingful method of determining and evaluating protein folding conditions, we have designed a new fractional factorial protein folding screen. The screen includes 12 factors shown by previous experiments to enhance protein foldi ng and it incorporates the 12 factors into 16 different folding conditions. By examining a 1/256(th) fraction of the full factorial, multiple folding conditions were determined for the ligand binding domains from glutamate an d kainate receptors, and for lysozyme and carbonic anhydrase B. The impact of each factor on the formation of biologically active material was estimat ed by calculating factor main effects. Factors and corresponding levels suc h as pH (8.5) and L-arginine (0.5 M) consistently had a positive effect on protein folding, whereas detergent (0.3 mM lauryl maltoside) and nonpolar a dditive (0.4 M sucrose) were detrimental to the folding of these four prote ins. One of the 16 conditions yielded the most folded material for three ou t of the four proteins. Our results suggest that this protein folding scree n will be generally useful in determining whether other proteins will fold in vitro and, if so, what factors are important. Furthermore, fractional fa ctorial folding screens are well suited to the evaluation of previously unt ested factors on protein folding.