Abstract
A simple capillary electrophoresis mobility shift assay (CEMSA), with no gel and uncoated capillaries, for the accurate determination of protein–DNA affinities free in solution was applied to constructs of the MyoD/E47 DNA-binding proteins. The determined affinities are compared to those obtained by EMSA. MyoD-E47 covalent heterodimer binds DNA more tightly ( K d =1.8 n M) than MyoD ( K d =14.2 n M) or E47 ( K d =11.5 n M) covalent homodimers. The effect of non-specific DNA on binding affinities was more important than salt concentration in the MyoD/E47 series. Application of this method to the MyoD/E47 system demonstrates the generality of our CEMSA.
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