Mechanisms controlling the temperature-dependent binding of proteins to poly(N-isopropylacrylamide) microgels.
Poly(N-isopropylacrylamide) (PNIPA) microgels may offer several advantages over PNIPA-modified surfaces when used as sorbents in temperature-sensitive chromatography. Yet, a full exploitation of these advantages requires a better understanding of the mechanisms controlling the separation process. As a model system, we have studied the binding of three proteins (bovine serum albumin (BSA), ovalbumin, and lysozyme) to PNIPA microgels. Binding experiments were conducted both below (25 degrees C) and above (37 degrees C) the volume phase transition temperature of the gel, T(c). The analysis of the binding isotherms has shown that although an average gel particle contained a larger amount of protein below the phase transition temperature, the concentration of the protein within the particle was higher above this temperature. These findings were attributed to changes in the binding loci due to temperature swings around T(c): whereas a sorption mechanism is dominant below this temperature, surface-adsorption was more important above it. A comparison between the three studied proteins has shown that below T(c) the binding increases with a decrease in the molecular weight. On the other hand, no significant difference in the bound protein amounts was observed above the phase transition temperature. Our results imply that, despite the increase in the gel's hydrophobicity above the phase transition temperature, the resolution in bioseparations based on PNIPA gels is not necessarily better above T(c).
Grabstain
Violet
V
Department of Chemical Engineering, Technion-Israel Institute of Technology, Haifa 32000, Israel.
Bianco-Peled
Havazelet
H
D000180:Acrylic Resins / Q000737:chemistry*; D002850:Chromatography, Gel / Q000379:methods*; D005782:Gels; D057927:Hydrophobic and Hydrophilic Interactions; D008422:Materials Testing; D008970:Molecular Weight; D009113:Muramidase / Q000032:analysis* / Q000737:chemistry*; D010047:Ovalbumin / Q000032:analysis* / Q000737:chemistry*; D044367:Phase Transition; D011485:Protein Binding; D011506:Proteins / Q000302:isolation & purification; D012710:Serum Albumin, Bovine / Q000032:analysis* / Q000737:chemistry*; D013696:Temperature
D000180:Acrylic Resins; D005782:Gels; D011506:Proteins; C052970:poly-N-isopropylacrylamide; D012710:Serum Albumin, Bovine; D010047:Ovalbumin; D009113:Muramidase