Molecular crowding creates an essential environment for the formation of stable G-quadruplexes in long double-stranded DNA.
Large numbers of guanine-rich sequences with potential to form G-quadruplexes have been identified in genomes of various organisms. Such sequences are constrained at both ends by long DNA duplex with a complementary strand in close proximity to compete for duplex formation. G-quadruplex/duplex competition in long double-stranded DNA has rarely been studied. In this work, we used DMS footprinting and gel electrophoresis to study G-quadruplex formation in long double-stranded DNA derived from human genome under both dilute and molecular crowding condition created by PEG. G-quadruplex formation was observed in the process of RNA transcription and after heat denaturation/renaturation under molecular crowding condition. Our results showed that the heat denaturation/renaturation treatment followed by gel electrophoresis could provide a simple method to quantitatively access the ability of G-quadruplex formation in long double-stranded DNA. The effect of K(+) and PEG concentration was investigated and we found that stable G-quadruplexes could only form under the crowding condition with PEG at concentrations near the physiological concentration of biomass in living cells. This observation reveals a physical basis for the formation of stable G-quadruplexes in genome and supports its presence under the in vivo molecular crowding condition.
Laboratory of Biochemistry and Biophysics, College of Life Sciences, Wuhan University, Wuhan, PR China.
Ke-wei
KW
Zheng
Zhao
Z
Chen
Yu-hua
YH
Hao
Zheng
Z
Tan
D004247:DNA / Q000737:chemistry*; D018983:DNA Footprinting; D004591:Electrophoresis, Polyacrylamide Gel; D054856:G-Quadruplexes; D006801:Humans; D011092:Polyethylene Glycols / Q000737:chemistry; D011188:Potassium / Q000737:chemistry; D013463:Sulfuric Acid Esters
D013463:Sulfuric Acid Esters; D011092:Polyethylene Glycols; D004247:DNA; C007482:dimethyl sulfate; D011188:Potassium