CONFORMATIONAL DIVERSITY AND CONFORMATIONAL TRANSITIONS OF A MONOCLONAL-ANTIBODY MONITORED BY CIRCULAR-DICHROISM AND CAPILLARY ELECTROPHORESIS

Four major isoforms of the BR96 antibody were separated by micellar el ectrokinetic capillary chromatography. Heat-induced reversible isoform interconversions were observed at 70 degrees C, and after extended in cubation at 80 degrees C, all species irreversibly transformed into a new single peak. In the presence of sodium dodecyl sulfate (1.0 mg/mL) , the isoform transformations occurred at lower temperatures without a ltering the separation pattern. Size exclusion chromatography analysis detected no aggregation at temperatures below 80 degrees C. Parallel circular dichroism measurements indicated significant conformational c hanges at 70-80 degrees C. The parallelism between isoform transformat ions and secondary structure changes allows consideration of CE-separa ted isoforms of BR96 antibody as conformers, an equilibrium between wh ich can be shifted by different physicochemical factors such as elevat ed temperatures and amphiphilic surfactants.