Ligand structure of the divinylsulfone-based T-gel.
The requirements for divinylsulfone (DVS)-based gels to act as thiophilic adsorbents, binding immunoglobulins in a salt-dependent manner have been examined. No differences in protein binding were observed for a DVS-activated gel reacted with mercaptoethanol (the T-gel), or for the same gel treated at high pH to hydrolyse the active groups and/or allow the formation of cross-links within the matrix, indicating that an O atom may be substituted for the thioether without affecting the thiophilic interactions. Extending the time of the activation reaction between DVS and the matrix results in increased amounts of sulfone attached to the gel, but decreased levels of active vinyl groups. When coupled to mercaptoethanol, these adsorbents bound more IgG than gels activated for shorter periods. This provides a convenient method to prepare thiophilic adsorbents of high capacity while minimising the amount of DVS used. The immobilised vinylsulfone must be linked to an electron donating atom for IgG to bind. When the vinyl was instead reduced with sodium borohydride, protein binding was decreased. No IgG bound to amine-coupled DVS-activated adsorbents, perhaps due to an overall positive charge on these gels at pH 7.4. The binding of human IgG to the adsorbents is dependent on ligand density, with little protein binding to gels having less than 16 mumol sulfone per ml. The binding increased with the ligand density above this level, with more than 25 mg IgG binding per ml to an adsorbent having 114 mumol sulfone per ml. The lack of binding at low ligand densities would be expected if the IgG must interact with two or more sulfone ligands to be retained on the adsorbent.
Scoble
J A
JA
School of Biochemistry, La Trobe University, Bundoora, Victoria, Australia. J.Scoble@latrobe.edu.au
Scopes
R K
RK
D001894:Borohydrides; D003432:Cross-Linking Reagents; D005782:Gels / Q000138:chemical synthesis / Q000737:chemistry* / Q000378:metabolism; D006868:Hydrolysis; D007075:Immunoglobulin M / Q000378:metabolism; D008024:Ligands; D008623:Mercaptoethanol; D010084:Oxidation-Reduction; D011485:Protein Binding; D012685:Sepharose / Q000737:chemistry*; D013329:Structure-Activity Relationship; D013450:Sulfones
D001894:Borohydrides; D003432:Cross-Linking Reagents; D005782:Gels; D007075:Immunoglobulin M; D008024:Ligands; D013450:Sulfones; C009873:divinyl sulfone; D008623:Mercaptoethanol; D012685:Sepharose