Separation of isoforms of Serratia marcescens nuclease by capillary electrophoresis.

Abstract

Three S. marcescens nuclease isoforms differing mainly in charge (native nuclease with pI 6.8 and two minor isoforms with pI 7.3 and 7.4) were separated using several different modes of high-performance capillary electrophoresis. Separation of the isoforms by free solution capillary electrophoresis was unsatisfactory. Separation by micellar electrokinetic capillary chromatography was therefore investigated in detail and the method optimized with respect to pH and sodium dodecyl sulphate concentration; in addition, the effect of adding various substances to control dispersion and avoid analyte adsorption at the capillary wall was examined. Under optimal conditions there was almost complete baseline separation of the two isoforms with basic pI whereas there was only partial separation of the native form and the isoform with pI 7.4. With capillary isoelectric focusing there was complete baseline separation of the native nuclease and the other two isoforms.