Structural basis for DNA recognition and allosteric control of the retinoic acid receptors RAR–RXR

Abstract Retinoic acid receptors (RARs) as a functional heterodimer with retinoid X receptors (RXRs), bind a diverse series of RA-response elements (RAREs) in regulated genes. Among them, the non-canonical DR0 elements are bound by RXR–RAR with comparable affinities to DR5 elements but DR0 elements do not act transcriptionally as independent RAREs. In this work, we present structural insights for the recognition of DR5 and DR0 elements by RXR–RAR heterodimer using x-ray crystallography, small angle x-ray scattering, and hydrogen/deuterium exchange coupled to mass spectrometry. We solved the crystal structures of RXR–RAR DNA-binding domain in complex with the Rarb2 DR5 and RXR–RXR DNA-binding domain in complex with Hoxb13 DR0. While cooperative binding was observed on DR5, the two molecules bound non-cooperatively on DR0 on opposite sides of the DNA. In addition, our data unveil the structural organization and dynamics of the multi-domain RXR–RAR DNA complexes providing evidence for DNA-dependent allosteric communication between domains. Differential binding modes between DR0 and DR5 were observed leading to differences in conformation and structural dynamics of the multi-domain RXR–RAR DNA complexes. These results reveal that the topological organization of the RAR binding element confer regulatory information by modulating the overall topology and structural dynamics of the RXR–RAR heterodimers.

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Purification, crystallization and X-ray diffraction analysis of the DNA-binding domain of human heat-shock factor 2

Acta Crystallographica Section F Structural Biology Communications ◽

10.1107/s2053230x16003599 ◽

2016 ◽

Vol 72 (4) ◽

pp. 294-299 ◽

Cited By ~ 3

Author(s):

Han Feng ◽

Wei Liu ◽

Da-Cheng Wang

Keyword(s):

Heat Shock ◽

Dna Binding ◽

Heat Shock Response ◽

Protein Family ◽

Binding Domain ◽

Structural Basis ◽

Dna Binding Domain ◽

X Ray Diffraction ◽

X Ray ◽

Shock Response

Cells respond to various proteotoxic stimuli and maintain protein homeostasis through a conserved mechanism called the heat-shock response, which is characterized by the enhanced synthesis of heat-shock proteins. This response is mediated by heat-shock factors (HSFs). Four genes encoding HSF1–HSF4 exist in the genome of mammals. In this protein family, HSF1 is the orthologue of the single HSF in lower eukaryotic organisms and is the major regulator of the heat-shock response, while HSF2, which shows low sequence homology to HSF1, serves as a developmental regulator. Increasing evidence has revealed biochemical properties and functional roles that are unique to HSF2, such as its DNA-binding preference and sumoylation patterns, which are distinct from those of HSF1. The structural basis for such differences, however, is poorly understood owing to the lack of available mammalian HSF structures. The N-terminal DNA-binding domain (DBD) is the most conserved functional module and is the only crystallizable domain in HSFs. To date, only HSF1 homologue structures from yeast and fruit fly have been determined. Along with extensive studies of the HSF family, more structural information, particularly from members with a remoter phylogenic relationship to the reported structures,e.g.HSF2, is needed in order to better understand the detailed mechanisms of HSF biology. In this work, the recombinant DBD (residues 7–112) from human HSF2 was produced inEscherichia coliand crystallized. An X-ray diffraction data set was collected to 1.32 Å resolution from a crystal belonging to space groupP212121with unit cell-parametersa= 65.66,b= 67.26,c= 93.25 Å. The data-evaluation statistics revealed good quality of the collected data, thus establishing a solid basis for the determination of the first structure at atomic resolution in this protein family.

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Structural basis of the p53 DNA binding domain and PUMA complex

Biochemical and Biophysical Research Communications ◽

10.1016/j.bbrc.2021.02.049 ◽

2021 ◽

Vol 548 ◽

pp. 39-46

Author(s):

Chang Woo Han ◽

Han Na Lee ◽

Mi Suk Jeong ◽

So Young Park ◽

Se Bok Jang

Keyword(s):

Dna Binding ◽

Binding Domain ◽

Structural Basis ◽

Dna Binding Domain

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Rgg protein structure–function and inhibition by cyclic peptide compounds

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1500357112 ◽

2015 ◽

Vol 112 (16) ◽

pp. 5177-5182 ◽

Cited By ~ 46

Author(s):

Vijay Parashar ◽

Chaitanya Aggarwal ◽

Michael J. Federle ◽

Matthew B. Neiditch

Keyword(s):

Dna Binding ◽

Structure Function ◽

Cyclosporin A ◽

Crystal Structures ◽

Disulfide Bond ◽

Binding Domain ◽

Dna Binding Domain ◽

X Ray ◽

Peptide Pheromone ◽

Dimerization Interface

Peptide pheromone cell–cell signaling (quorum sensing) regulates the expression of diverse developmental phenotypes (including virulence) in Firmicutes, which includes common human pathogens, e.g.,Streptococcus pyogenesandStreptococcus pneumoniae. Cytoplasmic transcription factors known as “Rgg proteins” are peptide pheromone receptors ubiquitous in Firmicutes. Here we present X-ray crystal structures of aStreptococcusRgg protein alone and in complex with a tight-binding signaling antagonist, the cyclic undecapeptide cyclosporin A. To our knowledge, these represent the first Rgg protein X-ray crystal structures. Based on the results of extensive structure–function analysis, we reveal the peptide pheromone-binding site and the mechanism by which cyclosporin A inhibits activation of the peptide pheromone receptor. Guided by the Rgg–cyclosporin A complex structure, we predicted that the nonimmunosuppressive cyclosporin A analog valspodar would inhibit Rgg activation. Indeed, we found that, like cyclosporin A, valspodar inhibits peptide pheromone activation of conserved Rgg proteins in medically relevantStreptococcusspecies. Finally, the crystal structures presented here revealed that the Rgg protein DNA-binding domains are covalently linked across their dimerization interface by a disulfide bond formed by a highly conserved cysteine. The DNA-binding domain dimerization interface observed in our structures is essentially identical to the interfaces previously described for other members of the XRE DNA-binding domain family, but the presence of an intermolecular disulfide bond buried in this interface appears to be unique. We hypothesize that this disulfide bond may, under the right conditions, affect Rgg monomer–dimer equilibrium, stabilize Rgg conformation, or serve as a redox-sensitive switch.

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