Recombinant proteins biologic activity

The significance of protein oxidation became paramount with the advent of recombinant protein biologies used as human therapeutics. Careful characterization of protein stability is essential to maintaining the efficacy of protein pharmaceuticals. If even a single side chain amino acid residue becomes oxidized, then a protein therapeutic may not have the same activity in vivo as the unmodified protein. 

Kumada Y, Shiritani Y, Hamasaki K, Ohse T, Kishimoto M (2009) High biological activity of a recombinant protein immobilized onto polystyrene. Biotechnol J 4 1178-1189... 

Studies have shown that plants can make biologically active recombinant proteins through both transgenic and transient expression approaches. Although the plant post-translational machinery is similar to that of mammalian cells, there are some notable differences, e.g. differences in glycosylation, particularly the absence of sia-lation, which may impact the activity of certain proteins. The absence of mammalian enzymes may prevent complex maturation processes that are critical for the biological activity of proteins such as insulin. Fortunately these shortcomings affect the activity of only a limited number of proteins. 

Recombinant proteins with unique properties can potentially generate new markets and penetrate into existing markets if they can be supplied on a large scale. An ideal system would produce the safest biologically active material at the lowest cost, and would be used in combination with an inexpensive and simple purification process. So far, there have been several examples of the high-yield production of recombinant proteins in transgenic crop plants, mainly in the area of molecular medicines such as antibodies, enzymes and vaccines [45, 48-50]. Modern agricultural practices offer... 

If a recombinant protein is developed and expressed for research purposes, the most labor- and cost-intensive part of the project normally lies with the upstream tasks, i.e. cloning, expression vector design, sequencing, transformation and selection. Once a suitable expresser strain or plant line is available, it is often sufficient to visualize expression of the recombinant protein, verify its activity, study its biological... 

Mammalian cell suspension cultures are the preferred choice for large-scale recombinant protein production in stirred-tank bioreactors. The most widely used systems are Chinese hamster ovary (CHO) cells and the murine myeloma fines NSO and SP2/0. In half of the biological license approvals from 1996-2000, CHO cells were used for the production of monoclonal antibodies and other recombinant glycosylated proteins, including tPA (tissue plasminogen activator) and an IgGl fusion with the tumor necrosis factor (TNF) receptor, the latter marketed as Enbrel [7]. 

Most interferons have now been produced in a variety of expression systems, including E. coli, fungi, yeast and some mammalian cell lines, such as CHO cell lines and monkey kidney cell lines. Most interferons currently in medical use are recombinant human (rh) products produced in E. coli. E. coli s inability to carry out post-translational modifications is irrelevant in most instances, as the majority of human IFN-as, as well as IFN- 3, are not normally glycosylated. Whereas IFN-y is glycosylated, the E. coli-derived unglycosylated form displays a biological activity similiar to the native human protein. 

A second unanswered concern is whether the antibody induced by the recombinant protein has any discernible health effect. Other than some reports of neutralization of biological activity, little pathology has been attributed to the presence of antibodies in patients given recombinant protein therapy. It should also be noted that the question of antibody specificity has not been well studied, so that it is entirely conceivable that autoimmune pathology or even an anaphylaxis response could be induced. Equally important is the concern that induced antibody might neutralize the endogenous hormone or protein that it is intended to replace or supplement. 

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