Molecular charge and electrophoretic mobility in cetacean myoglobins of known sequence

Abstract

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.

Clonal dynamics ofEscherichia coli in its natural habitat

“…This is because differences in the rate of migration of protein variants are a direct consequence of amino acid substitutions that affect net electrostatic charge (Selander et al 1986). Electrophoretic studies of protein variants of known amino acid sequence have demonstrated that this method detects most nucleotide substitutions resulting in amino acid replacements (Ramshaw et al 1979;McLellan 1984). However, the allelic variation revealed electrophoretically underestimates the total genetic variation at a locus because of silent nucleotide substitutions (Kreitman 1983) and amino acid replacements that do not alter mobility under standard conditions (Coyne 1982).…”

The separation of whale myoglobins with two-dimensional electrophoresis

“…The one-dimensional procedure separated all of the myoglobins studied here, but under only one of the five electrophoretic conditions used (McLellan, 1984). Under the other four buffer conditions the five myoglobins resolved into only three or four mobility classes.…”

“…The five rnyoglobins (Table I) (McLellan, 1984). The lyophilized protein was dissolved at a concentration of 0.125 mg/rnl in a 9 M urea, 2% Nonidet P-40 detergent, 2% mercaptoethanol, and 2% pH 9-11 arnpholyte (LKB) mixture.…”

Assignment of uroporphyrinogen decarboxylase <i>(UROD)</i> to the pter^→p21 region of human chromosome 1

“…1978). It is possible to have small sequence differences between proteins which result in charge differences at some pH values but not at others (McLellan, 1984). Relative electrophoretic mobility can be maximized near the isoelectric point, where the difference in charge between two proteins will be greatest, as the total charge o f each is small.…”