DNA-binding affinities of MyoD and E47 homo- and hetero-dimers by capillary electrophoresis mobility shift assay

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...

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Bibliographic Details
Published in:Journal of Chromatography A 1999-11, Vol.862 (2), p.231-236
Main Authors: Foulds, Glenn J, Etzkorn, Felicia A
Format: Article
Language:English
Subjects:
Algorithms
Biological and medical sciences
DNA
DNA - chemistry
DNA-Binding Proteins - chemistry
Electrophoresis, Capillary
Fundamental and applied biological sciences. Psychology
Gels
Helix-Loop-Helix Motifs
Interactions. Associations
Intermolecular phenomena
Molecular biophysics
MyoD Protein - chemistry
MyoD-E47 protein
TCF Transcription Factors
Transcription Factor 7-Like 1 Protein
Transcription Factors
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Summary: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.
ISSN:0021-9673
DOI:10.1016/S0021-9673(99)00923-1