Detailed kinetic analysis of the interaction between the FOXO4-DNA-binding domain and DNA.

作者：Vacha Petr; Zuskova Iva; Bumba Ladislav; Herman Petr; Vecer Jaroslav; Obsilova Veronika; Obsil Tomas

来源：Biophysical chemistry, 2013, 184: 68-78.

DOI：10.1016/j.bpc.2013.09.002

• Binding kinetics
• DNA-binding domain
• FOXO4 forkhead transcription factor
• Segmental dynamics
• Surface plasmon resonance
• Time-resolved fluorescence

The FOXO forkhead transcription factors are potent transcriptional activators involved in a wide range of key biological processes. In this work, the real-time kinetics of the interaction between the FOXO4-DNA binding domain (FOXO4-DBD) and the DNA was studied by using surface plasmon resonance (SPR). SPR analysis revealed that the interaction between FOXO4-DBD and the double stranded DNA containing either the insulin-responsive or the Daf-16 family member-binding element is preferably described by using a conformational change model which suggests a structural change of FOXO4-DBD upon binding to the DNA. This was further confirmed by using the time-resolved tryptophan fluorescence anisotropy decay measurements which revealed profound reduction of segmental dynamics of FOXO4-DBD upon the complex formation. Alanine scanning of amino acid residues engaged in polar contacts with the DNA showed that certain non-specific contacts with the DNA backbone are very important for both the binding affinity and the binding specificity of FOXO4-DBD.

• 单位
  Charles University