Microscopic understanding of the conformational features of a protein–DNA complex

AbstractThe interactions between protein-DNA are essential for various biological activities. In this review, we provide an overview of protein-DNA interactions that emphasizes the importance of dynamical aspects. We divide protein-DNA interactions into two categories: nonspecific and specific and both the categories would be discussed highlighting some of our relevant work. In the case of nonspecific protein-DNA interaction, solvation studies (picosecond and femtosecond-resolved) explore the role environmental dynamics and change in the micropolarity around DNA molecules upon complexation with histone protein (H1). While exploring the specific protein-DNA interaction at λ-repressor-operator sites interaction, particularly OR1 and OR2, it was observed that the interfacial water dynamics is minimally perturbed upon interaction with DNA, suggesting the labile interface in the protein-DNA complex. Förster resonance energy transfer (FRET) study revealed that the structure of the protein is more compact in repressor-OR2 complex than in the repressor-OR1 complex. Fluorescence anisotropy studies indicated enhanced flexibility of the C-terminal domain of the repressor at fast timescales after complex formation with OR1. The enhanced flexibility and different conformation of the C-terminal domain of the repressor upon complexation with OR1 DNA compared to OR2 DNA were found to have pronounced effect on the rate of photoinduced electron transfer.

Download Full-text

Mediators of activation of fushi tarazu gene transcription by BmFTZ-F1.

Molecular and Cellular Biology ◽

10.1128/mcb.14.5.3013 ◽

1994 ◽

Vol 14 (5) ◽

pp. 3013-3021 ◽

Cited By ~ 48

Author(s):

F Q Li ◽

H Ueda ◽

S Hirose

Keyword(s):

Transcriptional Activation ◽

Direct Contact ◽

In Vitro Transcription ◽

Tata Binding Protein ◽

Dna Interactions ◽

Fushi Tarazu ◽

Eukaryotic Sequence ◽

Protein Dna Interactions ◽

Dna Complex

Transcriptional activation by many eukaryotic sequence-specific regulators appears to be mediated through transcription factors which do not directly bind to DNA. BmFTZ-F1 is a silkworm counterpart of FTZ-F1, a sequence-specific activator of the fushi tarazu gene in Drosophila melanogaster. We report here the isolation of 18- and 22-kDa polypeptides termed MBF1 and MBF2, respectively, that form a heterodimer and mediate activation of in vitro transcription from the fushi tarazu promoter by BmFTZ-F1. Neither MBF1, MBF2, nor a combination of them binds to DNA. MBF1 interacts with BmFTZ-F1 and stabilizes the BmFTZ-F1-DNA complex. MBF1 also makes direct contact with TATA-binding protein (TBP). Both MBF1 and MBF2 are necessary to form a complex between BmFTZ-F1 and TBP. We propose a model in which MBF1 and MBF2 form a bridge between BmFTZ-F1 and TBP and mediate transactivation by stabilizing the protein-DNA interactions.

Download Full-text

Mediators of activation of fushi tarazu gene transcription by BmFTZ-F1

Molecular and Cellular Biology ◽

10.1128/mcb.14.5.3013-3021.1994 ◽

1994 ◽

Vol 14 (5) ◽

pp. 3013-3021

Author(s):

F Q Li ◽

H Ueda ◽

S Hirose

Keyword(s):

Transcriptional Activation ◽

Direct Contact ◽

In Vitro Transcription ◽

Tata Binding Protein ◽

Dna Interactions ◽

Fushi Tarazu ◽

Eukaryotic Sequence ◽

Protein Dna Interactions ◽

Dna Complex

Transcriptional activation by many eukaryotic sequence-specific regulators appears to be mediated through transcription factors which do not directly bind to DNA. BmFTZ-F1 is a silkworm counterpart of FTZ-F1, a sequence-specific activator of the fushi tarazu gene in Drosophila melanogaster. We report here the isolation of 18- and 22-kDa polypeptides termed MBF1 and MBF2, respectively, that form a heterodimer and mediate activation of in vitro transcription from the fushi tarazu promoter by BmFTZ-F1. Neither MBF1, MBF2, nor a combination of them binds to DNA. MBF1 interacts with BmFTZ-F1 and stabilizes the BmFTZ-F1-DNA complex. MBF1 also makes direct contact with TATA-binding protein (TBP). Both MBF1 and MBF2 are necessary to form a complex between BmFTZ-F1 and TBP. We propose a model in which MBF1 and MBF2 form a bridge between BmFTZ-F1 and TBP and mediate transactivation by stabilizing the protein-DNA interactions.

Download Full-text

ChIP-AP – An Integrated ChIP-Seq Analysis Pipeline

10.1101/2021.04.18.440382 ◽

2021 ◽

Author(s):

Jeremiah Suryatenggara ◽

Kol Jia Yong ◽

Danielle E. Tenen ◽

Daniel G. Tenen ◽

Mahmoud A. Bassal

Keyword(s):

Bioinformatics Analysis ◽

Peak Calling ◽

Dna Interactions ◽

Analysis Pipeline ◽

Multiple Peak ◽

Single File ◽

Protein Dna Interactions ◽

Dna Complex ◽

Peak Caller ◽

Peak Set

AbstractChIP-Seq is a technique used to analyse protein-DNA interactions. The protein-DNA complex is pulled down using a protein antibody, after which sequencing and analysis of the bound DNA fragments is performed. A key bioinformatics analysis step is “peak” calling - identifying regions of enrichment. Benchmarking studies have consistently shown that no optimal peak caller exists. Peak callers have distinct selectivity and specificity characteristics which are often not additive and seldom completely overlap in many scenarios. In the absence of a universal peak caller, we rationalized one ought to utilize multiple peak-callers to 1) gauge peak confidence as determined through detection by multiple algorithms, and 2) more thoroughly survey the protein-bound landscape by capturing peaks not detected by individual peak callers owing to algorithmic limitations and biases. We therefore developed an integrated ChIP-Seq Analysis Pipeline (ChIP-AP) which performs all analysis steps from raw fastq files to final result, and utilizes four commonly used peak callers to more thoroughly and comprehensively analyse datasets. Results are integrated and presented in a single file enabling users to apply selectivity and sensitivity thresholds to select the consensus peak set, the union peak set, or any sub-set in-between to more confidently and comprehensively explore the protein-bound landscape. (https://github.com/JSuryatenggara/ChIP-AP).

Download Full-text

Protein–DNA interactions: a novel approach to improve the fluorescence stability of DNA/Ag nanoclusters

Nanoscale ◽

10.1039/c4nr04889j ◽

2015 ◽

Vol 7 (4) ◽

pp. 1296-1300 ◽

Cited By ~ 7

Author(s):

Zhixue Zhou ◽

Shaojun Dong

Keyword(s):

Dna Binding ◽

Dna Interactions ◽

P Protein ◽

Novel Approach ◽

Protein Dna Interactions ◽

Ag Nanoclusters

Protein–DNA binding events can greatly improve the fluorescence stability of DNA-templated Ag nanoclusters.

Download Full-text

Halophilic adaptation of protein–DNA interactions

Biochemical Society Transactions ◽

10.1042/bst0310677 ◽

2003 ◽

Vol 31 (3) ◽

pp. 677-680 ◽

Cited By ~ 24

Author(s):

S. Bergqvist ◽

M.A. Williams ◽

R. O'Brien ◽

J.E. Ladbury

Keyword(s):

Dna Interaction ◽

Dna Interactions ◽

Evolutionary Time ◽

Homologous Proteins ◽

Binding Characteristics ◽

Dna Binding Site ◽

Protein Dna Interactions ◽

Tata Box Binding Protein ◽

Dna Complex ◽

Halophilic Adaptation

Pyrococcus woesei (Pw) is an archaeal organism adapted to living in conditions of elevated salt and temperature. Thermodynamic data reveal that the interaction between the TATA-box-binding protein (TBP) from this organism and DNA has an entirely different character to the same interaction in mesophilic counterparts. In the case of the PwTBP, the affinity of its interaction with DNA increases with increasing salt concentration. The opposite effect is observed in all known mesophilic protein–DNA interactions. The halophilic behaviour can be attributed to sequestration of cations into the protein–DNA complex. By mutating residues in the PwTBP DNA-binding site, potential sites of cation interaction can be removed. These mutations have a significant effect on the binding characteristics, and the halophilic nature of the PwTBP–DNA interaction can be reversed, and made to resemble that of a mesophile, in just three mutations. The genes of functionally homologous proteins in organisms existing in different environments show that adaptation is most often accompanied by mutation of an existing protein. However, the importance of any individual residue to a phenotypic characteristic is usually difficult to assess amongst the multitude of changes that occur over evolutionary time. Since the halophilic nature of this protein can be attributed to only three mutations, this reveals that the important phenotype of halophilicity could be rapidly acquired in evolutionary time.

Download Full-text

Assembly of the Hap2p/Hap3p/Hap4p/Hap5p-DNA Complex in Saccharomyces cerevisiae

Eukaryotic Cell ◽

10.1128/ec.4.11.1829-1839.2005 ◽

2005 ◽

Vol 4 (11) ◽

pp. 1829-1839 ◽

Cited By ~ 52

Author(s):

David S. McNabb ◽

Inés Pinto

Keyword(s):

Saccharomyces Cerevisiae ◽

Transcription Factor ◽

Dna Binding ◽

Transcriptional Activation ◽

Target Genes ◽

Dna Interactions ◽

Activation Domain ◽

Binding Factor ◽

Protein Dna Interactions ◽

Dna Complex

ABSTRACT The CCAAT-binding factor (CBF) is an evolutionarily conserved multimeric transcriptional activator in eukaryotes. In Saccharomyces cerevisiae, the CCAAT-binding factor is composed of four subunits, termed Hap2p, Hap3p, Hap4p, and Hap5p. The Hap2p/Hap3p/Hap5p heterotrimer is the DNA-binding component of the complex that binds to the consensus 5′-CCAAT-3′ sequence in the promoter of target genes. The Hap4p subunit contains the transcriptional activation domain necessary for stimulating transcription after interacting with Hap2p/Hap3p/Hap5p. In this report, we demonstrate that Hap2p, Hap3p, and Hap5p assemble via a one-step pathway requiring all three subunits simultaneously, as opposed to the mammalian CCAAT-binding factor which has been shown to assemble via a two-step pathway with CBF-A (Hap3p homolog) and CBF-C (Hap5p homolog) forming a stable dimer before CBF-B (Hap2p homolog) can interact. We have also found that the interaction of Hap4p with Hap2p/Hap3p/Hap5p requires DNA binding as a prerequisite. To further understand the protein-protein and protein-DNA interactions of this transcription factor, we identified the minimal domain of Hap4p necessary for interaction with the Hap2p/Hap3p/Hap5p-DNA complex, and we demonstrate that this domain is sufficient to complement the respiratory deficiency of a hap4Δ mutant and activate transcription when fused with the VP16 activation domain. These studies provide a further understanding of the assembly of the yeast CCAAT-binding factor at target promoters and raise a number of questions concerning the protein-protein and protein-DNA interactions of this multisubunit transcription factor.

Download Full-text