A new approach to examine conformational changes occurring upon binding of ligand by biomolecules.
Liquid-liquid partition chromatography in an aqueous poly(ethylene glycol)/dextran two-phase system (LLPC) is shown to be a quick and sensitive method for detecting conformational changes occurring upon binding of ligands by biospecific molecules. Two groups of well-characterized proteins, enzymes and monoclonal antibodies, were employed. As an example, LLPC demonstrated that isoforms of lactate dehydrogenase as well as of hexokinase existed in a ligand-dependent equilibrium between two forms and that conformational changes occurred when monoclonal antibodies bound haptens. We also demonstrate that the method could be used to detect and separate subfractions in preparations of unliganded proteins that appeared to be homogeneous when analysed by other techniques.
Wingren
C
C
Department of Biochemistry, University of Lund, Sweden.
Hansson
U B
UB
D000906:Antibodies / Q000737:chemistry; D000937:Antigen-Antibody Reactions; D002853:Chromatography, Liquid / Q000379:methods*; D003911:Dextrans; D006241:Haptens; D006593:Hexokinase / Q000737:chemistry; D066298:In Vitro Techniques; D007770:L-Lactate Dehydrogenase / Q000737:chemistry; D008024:Ligands; D008291:Malate Dehydrogenase / Q000737:chemistry; D011092:Polyethylene Glycols; D011485:Protein Binding; D011487:Protein Conformation
D000906:Antibodies; D003911:Dextrans; D006241:Haptens; D008024:Ligands; D011092:Polyethylene Glycols; D007770:L-Lactate Dehydrogenase; D008291:Malate Dehydrogenase; D006593:Hexokinase