Fast detection of quadruplex structure in DNA by the intrinsic fluorescence of a single-stranded DNA binding protein.
Single-stranded guanine-rich (G-rich) DNA can fold into a four-stranded G-quadruplex structure and such structures are implicated in important biological processes and therapeutic applications. So far, bioinformatic analysis has identified up to several hundred thousand of putative quadruplex sequences in the genome of human and other animal. Given such a large number of sequences, a fast assay would be desired to experimentally verify the structure of these sequences. Here we describe a method that identifies the quadruplex structure by a single-stranded DNA binding protein from a thermoautotrophic archaeon. This protein binds single-stranded DNA in the unfolded, but not in the folded form. Upon binding to DNA, its fluorescence can be quenched by up to 70%. Formation of quadruplex greatly reduces fluorescence quenching in a K+-dependent manner. This structure-dependent quenching provides simple and fast detection of quadruplex in DNA at low concentration without DNA labelling.
Zhuang
Xin-ying
XY
Laboratory of Biochemistry and Biophysics, College of Life Sciences, Wuhan University, Wuhan 430072, PR China.
Tang
Jun
J
Hao
Yu-hua
YH
Tan
Zheng
Z
D004247:DNA / Q000032:analysis* / Q000737:chemistry* / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D005453:Fluorescence; D054856:G-Quadruplexes; D006801:Humans; D006863:Hydrogen-Ion Concentration; D017017:Methanococcus / Q000737:chemistry*; D009841:Oligonucleotides / Q000032:analysis / Q000737:chemistry; D011188:Potassium / Q000494:pharmacology; D016615:Telomere / Q000737:chemistry
D004268:DNA-Binding Proteins; D009841:Oligonucleotides; D004247:DNA; D011188:Potassium