Effect of heat and sodium dodecyl sulfate on solubilization of proteins before two-dimensional polyacrylamide gel electrophoresis.

To solubilize biological samples, sodium dodecyl sulfate (SDS) frequently is added and the mixture heated at 70-100 degrees C. However, two-dimensional polyacrylamide gel electrophoresis of a single protein after SDS treatment has not been reported. When rabbit-muscle creatine kinase was so run, we saw considerable difference in the gel staining pattern for the heated and nonheated enzyme dissolved in the SDS solution. After heating for 10 min at 95 degrees C the number of silver-stained spots apparent increased, and staining of several spots intensified. After 60 min, most of the discrete spots disappeared. Evidently the peptide backbone had been hydrolyzed. When the enzyme was simply left at room temperature for four days, the effects were similar. Appearance of new spots and loss of spots apparently are caused by heating alone but are intensified by SDS. Experiments with human serum albumin yielded similar results.