Electrophoretic Mobility Shift Assay and Dimethyl Sulfate Footprinting for Characterization of G-Quadruplexes and G-Quadruplex-Protein Complexes

Buket Onel, Guanhui Wu, Daekyu Sun, Clement Lin, Danzhou Yang

Research output: Chapter in Book/Report/Conference proceedingChapter

Abstract

DNA G-quadruplexes are globular nucleic acid secondary structures which occur throughout the human genome under physiological conditions. There is accumulating evidence supporting G-quadruplex involvement in a number of important aspects of genome functions, including transcription, replication, and genomic stability, and that protein and enzyme recognition of G-quadruplexes may represent a key event to regulate physiological or pathological pathways. Two important techniques to study G-quadruplexes and their protein interactions are the electrophoretic mobility shift assay (EMSA) and dimethyl sulfate (DMS) footprinting assay. EMSA, one of the most sensitive and robust methods for studying the DNA-protein interactions, can be used to determine the binding parameters and relative affinities of a protein for the G-quadruplex. DMS footprinting is a powerful assay for the initial characterization of G-quadruplexes, which can be used to deduce the guanine bases involved in the formation of G-tetrads under physiological salt conditions. DMS footprinting can also reveal important information in G-quadruplex-protein complexes on protein contacts and regional changes in DNA G-quadruplex upon protein binding. In this paper, we will provide a detailed protocol for the EMSA and DMS footprinting assays for characterization of G-quadruplexes and G-quadruplex-protein complexes. Expected outcomes and references to extensions of the method will be further discussed.

Original languageEnglish (US)
Title of host publicationMethods in Molecular Biology
PublisherHumana Press Inc.
Pages201-222
Number of pages22
DOIs
StatePublished - Jan 1 2019

Publication series

NameMethods in Molecular Biology
Volume2035
ISSN (Print)1064-3745
ISSN (Electronic)1940-6029

Keywords

  • DNA
  • Dimethyl sulfate (DMS) footprinting
  • Electrophoresis
  • Electrophoretic mobility shift assay (EMSA)
  • G-quadruplex
  • Protein

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics

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    Onel, B., Wu, G., Sun, D., Lin, C., & Yang, D. (2019). Electrophoretic Mobility Shift Assay and Dimethyl Sulfate Footprinting for Characterization of G-Quadruplexes and G-Quadruplex-Protein Complexes. In Methods in Molecular Biology (pp. 201-222). (Methods in Molecular Biology; Vol. 2035). Humana Press Inc.. https://doi.org/10.1007/978-1-4939-9666-7_11