Purification of Recombinant Plant-Made Proteins From Corn by Ultrafiltration

Separations Division
2009 AIChE Annual Meeting
AIChE Annual Meeting
November 10, 2009 - 7:00pm
The use of corn as an expression host is an attractive system for the production of plant-made proteins. The use of plants lowers upstream costs making downstream the significant production cost. For some proteins, crossflow ultrafiltration could be a lower cost alternative. The potential for doing so for protein of two different sizes was explored in this work. Membrane filtration performance of a large molecular weight protein, forms of which have been expressed in corn – triple-helical type 1 recombinant human (rh) collagen - was contrasted with the performance of a small protein, green fluorescent protein (GFP). Studies were conducted by spiking rhCollagen into endosperm and germ extracts of transgenic corn expressing GFP. Crossflow ultrafiltration parameters of membrane pore size, transmembrane pressure (TMP), crossflow rate, and pH were studied for their effect on permeate flux and protein sieving.

Ultrafiltration resulted in excellent purity and high yield of collagen for both the germ and endosperm extracts. Filtration using a 100 kDa molecular weight cut off (MWCO) membrane resulted in complete retention of collagen, whereas the 300 kDa MWCO membrane showed significant loss of collagen throughout the filtration. For the 100 kDa membrane, no significant difference in the sieving of either the host cell proteins, GFP or rhCollagen occurred under different conditions of crossflow rate and TMP; however, a significant increase in the permeate flux was observed at higher crossflow and TMP. Additionally, increasing the pH resulted in a significant decrease in the sieving of host cell proteins while pH had no effect on GFP sieving. Filtrations performed in concentration mode were able to achieve collagen concentrations as high as 1 mg/ml without any observed sieving. These results indicate that ultrafiltration can be used to obtain a high purity when separating large proteins from corn extracts.&'
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