Summary
Covalent attachment of polyethylene glycol (PEG) to proteins produces an increase in their partition coefficient (K) in PEG/dextran aqueous two-phase systems. At least for bovine serum albumin (BSA) and granulocyte-macrophage colony stimulating factor (GM-CSF) a linear relationship exists between log K of the conjugate and the number of PEG molecules attached to it (up to 20 PEG molecules for BSA and 3 for GM-CSF). The proportionality constant (i.e., the increment in log K per PEG molecule attached) is protein specific. For PEGGM-CSF conjugates, the proportionality constant increases with the concentration of the polymers (PEG and dextran up to 6%, w/w) in the two-phase system. A further increase in the polymer concentration up to 6.5% leads to a decrease in the proportionality constant. To explain these observations it is proposed that the interaction between the PEG-modified conjugate and the polymers in the phase system includes a positive interaction between the PEG attached to the PEG-modified conjugate and the PEG coils in the top phase, in addition to the excluded volume interactions generally accepted as the main interactions determining the partitioning of native proteins.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Preview
Unable to display preview. Download preview PDF.
References
H. Walter, D.E. Brooks, and D. Fisher (eds.). “Partitioning in Aqueous Two-Phase Systems, Theory, Methods, Uses and Applications to Biotechnology,” Academic Press, New York (1985).
P.A. Albertsson. “Partition of Cell Particles and Macromolecules,” John Wiley and Sons, New York (1986).
D. Fisher and I.A. Sutherland (eds.). “Separations Using Aqueous Phase Systems. Applications in Cell Biology and Biotechnology,” Plenum Press, New York and London (1989).
A. Gordes, J. Flossdorf, and M.R. Kula, Affinity partitioning: development of a mathematical model describing behavior of biomolecules in aqueous two-phase systems, Biotechnol. Bioengineer. 30:514 (1987).
J.M. Harris, E.C. Struck, M.G. Case, M.S. Paley, M. Yalpani, J.M. van Alstine, and D.E. Brooks, Synthesis and characterization of polyethylene glycol derivatives, J. Polym. Sci. 22:341 (1984).
K.A. Sharp, M. Yalpani, S.J. Howard, and D.E. Brooks, Synthesis and application of a poly(ethylene glycol)-antibody affinity ligand for cell separations in aqueous polymer two-phase systems, Anal. Biochem. 154:110 (1986).
G. Kopperschlager and G. Johansson, Affinity partitioning with polymer-bound Cibacron blue F3G-A for rapid, large-scale purification of phosphofructokinase from Baker’s yeast, Anal. Biochem. 124:117 (1982).
G. Johansson and M. Joelsson, Affinity partitioning of enzymes using dextran-bound procion yellow HE-3G. Influence of dye-ligand density, J. Chromatogr. 393:195 (1987).
N.L. Abbott, D. Blankschtein, and T.A. Hatton, On protein partitioning in two-phase aqueous polymer systems, Bioseparation 1:191 (1990).
D. Forciniti and C.K. Hall, Theoretical treatment of aqueous two-phase extraction by using virial expansions. A preliminary report, in: “Downstream Processing and Bioseparations: Recovery and Purification of Biological Products,” ACS Symposium Series, Vol. 419, J.-F.P. Hamel, J.B. Hunter, and S.K. Sikdar, eds., American Chemical Society, Washington D.C. (1990).
N.L. Abbott, D. Blankschtein, and T.A. Hatton, Protein partitioning in two-phase aqueous polymer systems. 1. Novel physical pictures and a scaling-thermodynamic formulation, Macromolecules 24:4334 (1991).
C. Delgado, J.N. Patel, G.E. Francis, and D. Fisher, Coupling of poly(ethylene glycol) to albumin under very mild conditions by activation with tresyl chloride: characterization of the conjugate by partitioning in aqueous two-phase systems, Biotechnol. Appl. Biochem. 12:119 (1990).
S.J. Stocks, A.J. Jones, C.W. Ramey, and D.E. Brooks, A fluorometric assay of the degree of modification of protein primary amines with polyethylene glycol, Anal. Biochem. 154:232 (1986).
C. Delgado, F. Malik, B. Selisko, D. Fisher, and G.E. Francis, Quantitative analysis of polyethylene glycol (PEG) in PEG-modified proteins/cytokines by aqueous two-phase systems, J. Biochem. Biophys. Methods, in press (1994).
S. Sasakawa and H. Walter, Partition behavior of native proteins in aqueous dextran-poly(ethylene glycol)-phase systems, Biochemistry 11:2760 (1972).
C. Delgado, G.E. Francis, and D. Fisher, Uses and properties of PEG-linked proteins, in: “Critical Reviews in Therapeutic Drug Carrier Systems,” CRC Press, Boca Raton, Florida, (1992).
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 1995 Springer Science+Business Media New York
About this chapter
Cite this chapter
Delgado, C. (1995). Partitioning of Polyethylene Glycol(PEG)-Protein Conjugates in PEG/Dextran Aqueous Two-Phase Systems. In: Rogers, R.D., Eiteman, M.A. (eds) Aqueous Biphasic Separations. Springer, Boston, MA. https://doi.org/10.1007/978-1-4615-1953-9_14
Download citation
DOI: https://doi.org/10.1007/978-1-4615-1953-9_14
Publisher Name: Springer, Boston, MA
Print ISBN: 978-1-4613-5802-2
Online ISBN: 978-1-4615-1953-9
eBook Packages: Springer Book Archive