PEGylation purification

Reaction used at an appropriate point in purification train for conversion to liioactive forms (e.g. refold / oxidation, dimer formation, PEGylation)... 

The result of the inherent comparison of AChE and BChE that was completed in our laboratory revealed two very adept OP bioscavengers. The major difference noted in this study was an ineffectiveness of oxime-mediated reactivation of BChE relative to the accelerated rates seen with AChE (Geyer, Cerasoli, Lenz, and Mor, unpublished results). In the case of both enzymes, by utilizing purification protocols that were only mildly adapted from those used to purify serum BChE or recombinant ChEs we can assume that any associated purification costs will be similar to those incurred in other systems. Before this project is ready to transition into a clinical study, a further refinement of the PEGylation and other modifications that have been shown to enhance the circulatory profile of these plant-derived ChEs must be undertaken. 

Lee, E.K. and Lee, J.D., Solid-phase, N-terminus-specific, mono-PEGylation of recombinant inter-feron-alpha-2a Purification, characterization, and bioactivity, presented atint. Symp. on the Separation of Proteins, Peptides and Polynucleotides, Aachen, Eederal Repubhc of Germany, Oct 19-22, 2004. 

Xu, Y., Mehl, IT, Bakhtiar, R., et al. (2010) Immunoaffi-nity purification using anti-PEG antibody followed by two-dimensional liquid chromatography/tandem mass spectrometry tor the quantification of a PEGylated therapeutic peptide in human plasma. Analytical Chemistry, 82, 6877-6886. 

An elaborate but useful method for the preparation and use of 3 in the PEGylation of proteins that avoids intermediate purification steps has been published in the patent literature [37]. A noteworthy point is that only one purification step is performed after the conjugation reaction. 

R.M. Arduini, Z. Li, A. Rapoza, R. Gronke, D.M. Hess, D. Wen, K. Miatkowski, C. Coots, A. Kaffashan, N. Viseux, J. Delaney, B. Domon, C.N. Young, R. Boyton, L.L Chen, M. Betzenhauser, S. Miller, A. Gill, R.B. Pepinsky, P.S. Hochman, D.P. Baker, Expression, purification, and characterization of rat interferon-P, and preparation of an N-terminally PEGylated form with improved pharmacokinetic parameters. Protein Expres. Purif., 34 (2) 229-242,2004. 

X. Suo, X. Lu, T. Hu, G. Ma, Z. Su, A sohd-phase adsorption method for PEGylation of human serum albumin and staphylokinase preparation, purification and biochemical characterization. BiotechnoL Lett., 31 (8) 1191-1196,2009. 

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