物理化学学报 >> 2010, Vol. 26 >> Issue (04): 1099-1106.doi: 10.3866/PKU.WHXB20100439

生物物理化学 上一篇    下一篇

单分子荧光共振能量传递及系综生物物理表征人体肌细胞增强因子2D启动子形成的G-四链体

周文华, 应立明   

  1. Molecular Medicine, National Heart and Lung Institute, Imperial College London, London SW7 2AZ, United Kingdom; Chemical Biology Centre, Imperial College London, London SW7 2AZ, United Kingdom
  • 收稿日期:2009-12-03 修回日期:2010-02-03 发布日期:2010-04-02
  • 通讯作者: 应立明 E-mail:l.ying@imperial.ac.uk

Single Molecule Fluorescence Resonance Energy Transfer and Ensemble Biophysical Characterization of a G-quadruplex Formed in the Promoter of Human Myocyte Enhancer Factor 2D

ZHOU Wen-Hua, YING Li-Ming   

  1. Molecular Medicine, National Heart and Lung Institute, Imperial College London, London SW7 2AZ, United Kingdom; Chemical Biology Centre, Imperial College London, London SW7 2AZ, United Kingdom
  • Received:2009-12-03 Revised:2010-02-03 Published:2010-04-02
  • Contact: YING Li-Ming E-mail:l.ying@imperial.ac.uk

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摘要：

DNA G-quadruplexes (G4s) are believed to be involved in many biological processes, including the gene expression regulation and the maintenance of telomere. However, the kinetics of G4, in particular how G4 forms in the double-stranded genomic DNA is an underexplored issue. We found that a very stable G4 can form in the promoter of human myocyte enhancer factor 2D (MEF2D), the gene encodes a transcription factor belonging to the MEF2 (myocyte enhancer factor-2) family which regulates the response of heart to cardiac stress signals. Biophysical characterizations, in particular single molecule fluorescence resonance energy transfer (smFRET) measurements, suggest that this G4 is more stable than its duplex form in near physiological conditions. Two major G4 structures were identified based on single molecule conformational analysis on the end and internal labeled DNA systems. A free energy landscape for the G4 hybridization was established in light of the smFRET G4/duplex competition assay. In addition, the G4 hybridization and unfolding rate constants were obtained.

关键词: Fluorescence resonance energy transfer, DNA G-quadruplex, Myocyte enhancer factor 2D, Unfolding kinetics, Single molecule

Abstract:

DNA G-quadruplexes (G4s) are believed to be involved in many biological processes, including the gene expression regulation and the maintenance of telomere. However, the kinetics of G4, in particular how G4 forms in the double-stranded genomic DNA is an underexplored issue. We found that a very stable G4 can form in the promoter of human myocyte enhancer factor 2D (MEF2D), the gene encodes a transcription factor belonging to the MEF2 (myocyte enhancer factor-2) family which regulates the response of heart to cardiac stress signals. Biophysical characterizations, in particular single molecule fluorescence resonance energy transfer (smFRET) measurements, suggest that this G4 is more stable than its duplex form in near physiological conditions. Two major G4 structures were identified based on single molecule conformational analysis on the end and internal labeled DNA systems. A free energy landscape for the G4 hybridization was established in light of the smFRET G4/duplex competition assay. In addition, the G4 hybridization and unfolding rate constants were obtained.

Key words: Fluorescence resonance energy transfer, DNA G-quadruplex, Myocyte enhancer factor 2D, Unfolding kinetics, Single molecule