Molecular charge and electrophoretic mobility in cetacean myoglobins of known sequence.

Fourteen myoglobins of known sequence were examined by polyacrylamide gel electrophoresis at five pH values. Gels at each pH divided the sequences into six to eight distinct classes, while the combination of the results of three gels at different pH levels distinguished 13 of 14, or 93%, of the sequences. The relative mobility of the myoglobins in the gels is significantly correlated with the charges of the proteins calculated from the pK values of the ionized groups. Major differences in mobility corresponded to expected differences in charge due to the amino acid substitutions between sequences. In addition to sequences differing in the total number of acidic and basic residues, those differing from each other in the total number of histidines were distinguished on low-pH gels. One pair of sequences differing by the exchange of lysine for arginine was separated on high-pH gels, as predicted from the differences in ionization of these two amino acids. On gels at pH 10.4, there was greater deviation of electrophoretic mobility from charge than on other gels, possibly due to the influence of amino acid substitutions in the neighborhood of lysine residues. Manipulation of the concentration and composition of the gels did not change the separation of the sequences from each other. Examination of myoglobins by gel electrophoresis at a wide range of pH values allows discrimination of nearly all amino acid substitutions and demonstrates the close relationship between titration and relative electrophoretic mobility.