Purification of proteins produced by biotechnological process.
Proteins maintain functions important to life. Faulty functioning or deficiency gives rise to pathological reactions. These proteins can now be produced, using the methods of recombinant DNA technology and administered to patients for replacement therapy. Many proteins as active ingredients are already available for use as immunomodulators, agents for tumour treatment, plasma proteins and hormones. They are in various stages of development, ranging from cloning of the producing cells to marketing of the finished products. Since the active substances are proteins synthesized by recombinant cells, their purification presents a particular challenge to protein chemists. Purification of recombinant DNA-derived proteins intended for human use is an essential part of the biotechnical process. It starts immediately after the fermentation of the host cell. The characteristics of the protein determine which microorganisms or cell cultures are used and this in turn defines the first purification step. The microorganisms are disrupted, and the insoluble protein, desposited in "inclusion bodies" has to be renatured, or the proteins secreted by cells and have to be concentrated. The subsequent strategy for purification of the protein does not depend on the fermentation process but is entirely determined by the physiochemical properties of the proteins. The goal of the first purification step is to isolate as fast and quantitatively as possible the recombinant protein from the culture filtrate, in order to minimize potential changes brought about by proteases or glycosidases. Immunoaffinity or ligand-affinity chromatography is used preferentially for this purpose. The concentration of protein and buffer changes are carried out by precipitation followed by reconstitution or, preferably, by dialysis and ultrafiltration/diafiltration.(ABSTRACT TRUNCATED AT 250 WORDS)
Werner
R G
RG
Dr. Karl Thomae GmbH, Bioengineering Techniques, Biberach an der Riss, Fed. Rep. of Germany.
Berthold
W
W
D055598:Chemical Phenomena; D002627:Chemistry, Physical; D006023:Glycoproteins / Q000302:isolation & purification; D011994:Recombinant Proteins / Q000302:isolation & purification*
D006023:Glycoproteins; D011994:Recombinant Proteins