title: Detailed kinetic analysis of the interaction between the FOXO4-DNA-binding domain and DNA.
abstract: 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.
authors.affiliation: Department of Physical and Macromolecular Chemistry, Faculty of Science, Charles University in Prague, 12843 Prague, Czech Republic.
authors.forename: Petr
authors.initials: P
authors.lastname: Vacha
authors.forename: Iva
authors.initials: I
authors.lastname: Zuskova
authors.forename: Ladislav
authors.initials: L
authors.lastname: Bumba
authors.forename: Petr
authors.initials: P
authors.lastname: Herman
authors.forename: Jaroslav
authors.initials: J
authors.lastname: Vecer
authors.forename: Veronika
authors.initials: V
authors.lastname: Obsilova
authors.forename: Tomas
authors.initials: T
authors.lastname: Obsil
mesh_terms: D001665:Binding Sites; D018797:Cell Cycle Proteins; D004247:DNA / Q000737:chemistry*; D051858:Forkhead Transcription Factors; D006801:Humans; D007700:Kinetics; D008958:Models, Molecular; D017434:Protein Structure, Tertiary; D020349:Surface Plasmon Resonance; D014157:Transcription Factors / Q000737:chemistry*
chemical_list: D018797:Cell Cycle Proteins; C108803:FOXO4 protein, human; D051858:Forkhead Transcription Factors; D014157:Transcription Factors; D004247:DNA
keywords: Binding kinetics; DNA-binding domain; FOXO4 forkhead transcription factor; Segmental dynamics; Surface plasmon resonance; Time-resolved fluorescence