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At least 469 records · Page 26Linked to original sources
PubMed2001

A unique beta-hairpin protruding from AAA+ ATPase domain of RuvB motor protein is involved in the interaction with RuvA DNA recognition protein for branch migration of Holliday junctions.

The Escherichia coli RuvB protein is a motor protein that forms a complex with RuvA and promotes branch migration of Holliday junctions during homologous recombination. This study describes the characteristics of two RuvB mutants, I148T and I150T, that do not promote branch migration in the presence of RuvA. These RuvB mutants hydrolyzed ATP and bound duplex DNA with the same efficiency as wild-type RuvB, but the mutants did not form a complex with RuvA and were defective in loading onto junction DNA in a RuvA-assisted manner. A recent crystallographic study revealed that Ile(148) and Ile(150) are in a unique beta-hairpin that protrudes from the AAA(+) ATPase domain of RuvB. We propose that this beta-hairpin interacts with hydrophobic residues in the mobile third domain of RuvA and that this interaction is vital for the RuvA-assisted loading of RuvB onto Holliday junction DNA.

Adenosine Triphosphatases↗
PubMed1985

Thermosensitivity of a DNA recognition site: activity of a truncated nutL antiterminator of coliphage lambda.

Antitermination is an important transcriptional control. In bacteriophage lambda, the presence of the nut antiterminators between the promoters and terminators results in relatively unhindered transcription when the lambda N gene product and necessary host factors are supplied. This antitermination system has been rendered thermosensitivity by modification of the nut site. A fragment of lambda DNA [74 base pairs (bp) in length]that contained the 17-bp nutL core sequence, but lacked the 8-bp boxA sequence, was cloned in a pp-N-tL1-galK plasmid between the pp promoter and gene N. This fragment mediated antitermination of transcription at 30 degrees C, as measured by assaying galK gene expression in Escherichia coli. At 42 degrees C, however, antitermination at the lambda tL1 terminator was abolished. Antitermination at 42 degrees C was restored by replacing the 74-bp nutL fragment with longer sequences containing both nutL and boxA or by cloning a synthetic boxA sequence ahead of the 74-bp nutL fragment. Thus, efficient antitermination required both boxA and the 17-bp nutL core, with the latter becoming conditionally defective when the boxA sequence was deleted.

Bacteriophage lambda↗
PubMed2001

Structural analyses of DNA recognition by the AML1/Runx-1 Runt domain and its allosteric control by CBFbeta.

The core binding factor (CBF) heterodimeric transcription factors comprised of AML/CBFA/PEBP2alpha/Runx and CBFbeta/PEBP2beta subunits are essential for differentiation of hematopoietic and bone cells, and their mutation is intimately related to the development of acute leukemias and cleidocranial dysplasia. Here, we present the crystal structures of the AML1/Runx-1/CBFalpha(Runt domain)-CBFbeta(core domain)-C/EBPbeta(bZip)-DNA, AML1/Runx-1/CBFalpha(Runt domain)-C/EBPbeta(bZip)-DNA, and AML1/Runx-1/CBFalpha(Runt domain)-DNA complexes. The hydrogen bonding network formed among CBFalpha(Runt domain) and CBFbeta, and CBFalpha(Runt domain) and DNA revealed the allosteric regulation mechanism of CBFalpha(Runt domain)-DNA binding by CBFbeta. The point mutations of CBFalpha related to the aforementioned diseases were also mapped and their effect on DNA binding is discussed.

Allosteric Regulation↗
PubMed1982

The operator-binding domain of lambda repressor: structure and DNA recognition.

The structure of the operator-binding domain of the lambda repressor has been determined at 3.2 A resolution. This domain contains an extended N-terminal arm and five alpha-helices. Model-building studies of the repressor-operator complex suggest that alpha-helices, especially the N-terminal parts of these helices, may provide a useful surface for protein-DNA interactions.

Amino Acid Sequence↗
PubMed1991

Purification of the DNA binding domain of herpes simplex virus type 1 immediate-early protein Vmw175 as a homodimer and extensive mutagenesis of its DNA recognition site.

The herpes simplex virus type 1 (HSV-1) Immediate-Early (IE) polypeptide Vmw175 is essential for the activation of transcription from viral early and late promoters. Vmw175 also reduces the activity of its own (IE-3) promoter in transfection assays. Both transactivation and repression mediated by Vmw175 require the integrity of a conserved domain of the polypeptide which has been shown to bind to specific DNA sequences. We have investigated the DNA sequence requirements for Vmw175 binding using a randomly mutated target. The results indicate that the binding site covers a region of 13 nucleotides divided into proximal and distal parts which are consistent with the consensus ATCGTNNNNNYSG. We have also expressed several different constructs encompassing the DNA binding domain of Vmw175 in bacteria, and obtained preparations of greater than 90% purity. The DNA binding domain is a dimer in solution, and binds DNA with a specificity similar to that of the intact protein, although the smallest DNA binding competent protein has a slightly reduced specificity.

Base Sequence↗
PubMed1991

Nucleotide sequence, messenger RNA stability, and DNA recognition elements of cys-14, the structural gene for sulfate permease II in Neurospora crassa.

The complete nucleotide sequence of the cys-14 gene which encodes sulfate permease II, a member of the sulfur regulatory circuit, is presented. The cys-14 gene contains four introns with consensus splice site sequences and is transcribed from four closely spaced initiation sites located approximately 20 bp upstream of the ATG initiation codon. The translated CYS14 protein is composed of 781 amino acids with a molecular weight of 87,037 and contains 12 potential hydrophobic membrane-spanning domains. cys-4 mRNA was found to turn over with a half-life of approximately 15 min, which presumably contributes to the regulation of sulfate permease II function. The cys-14 gene is highly expressed, but only in cells subject to sulfur limitation, and is turned on by the positive-acting CYS3 sulfur regulatory protein. Results are presented which show that CYS3 protein binds with higher affinity to DNA fragments which contain two or three tandem copies of a binding site sequence. Analyses of binding site specificity via mutated binding site elements showed that different regions of the partially symmetrical CYS3 binding site are important for recognition by the CYS3 regulatory protein.

Amino Acid Sequence↗
PubMed1988

The DNA recognition site for the dioxin-Ah receptor complex. Nucleotide sequence and functional analysis.

The environmental contaminant 2,3,7,8-tetra-chlorodibenzo-p-dioxin produces its biological effects by binding to an intracellular protein (the Ah receptor). The dioxin-Ah receptor complex activates cytochrome P1-450 gene transcription by interacting with dioxin-responsive enhancers. Here, we report that the dioxin-Ah receptor complex binds to DNA containing the "core" sequence 5'-TA/TGCGTG-3', which is present in each of three receptor-dependent enhancers. Functional analyses indicate that binding of the liganded Ah receptor to the core sequence fails to generate an active enhancer and that nucleotides flanking the core sequence must contribute to enhancer function and dioxin action.

Animals↗
PubMed1986

Role of thymidine residues in DNA recognition by the EcoRI and EcoRV restriction endonucleases.

We have synthesized a series of oligonucleotides containing the EcoRI (GAATTC) or EcoRV (GATATC) recognition site within which or adjacent to which thymidine was substituted by uridine or derivatives of uridine. The effects of these substitutions on the rate of the EcoRI and EcoRV catalyzed cleavage reaction were investigated. Our results show that most of the substitutions within the site are quite well tolerated by EcoRI, not, however, by EcoRV. We conclude that the thymin residues most likely are not directly involved in the recognition process of the EcoRI reaction. In contrast, they are major points of contact, between substrate and enzyme in the EcoRV reaction. The effects of substitutions in the position adjacent to the recognition site is also markedly different for EcoRI and EcoRV. Here, EcoRI seems to be considerably more selective than EcoRV.

Base Sequence↗
PubMed2002

Influence of a terminal formamido group on the sequence recognition of DNA by polyamides.

Pyrrole (Py)-imidazole (Im)-containing polyamides bind in the minor groove of DNA and can recognize specific sequences through a stacked antiparallel dimer. It has been proposed that there are two different low energy ways to form the stacked dimer and that these are sensitive to the presence of a terminal formamido group: (i) a fully overlapped stacking mode in which the N-terminal heterocycles of the dimer stack on the amide groups between the two heterocycles at the C-terminal and (ii) a staggered stacking mode in which the N-terminal heterocycles are shifted by approximately one unit in the C-terminal direction (Structure 1997, 5, 1033-1046). Two different DNA sequences will be recognized by the same polyamide stacked in these two different modes. Despite the importance of polyamides as sequence specific DNA recognition agents, these stacking possibilities have not been systematically explored. As part of a program to develop agents that can recognize mismatched base pairs in DNA, a set of four polyamide trimers with and without terminal formamido groups was synthesized, and their interactions with predicted DNA recognition sequences in the two different stacking modes were evaluated. Experimental difficulties in monitoring DNA complex formation with polyamides were overcome by using surface plasmon resonance (SPR) detection of the binding to immobilized DNA hairpin duplexes. Both equilibrium and kinetic results from SPR show that a terminal formamido group has a pronounced effect on the affinity, sequence specificity, and rates of DNA-dimer complex formation. The formamido polyamides bind preferentially in the staggered stacking mode, while the unsubstituted analogues bind in the overlapped mode. Affinities for cognate DNA sequences increase by a factor of around 100 when a terminal formamido is added to a polyamide, and the preferred sequences recognized are also different. Both the association and the dissociation rates are slower for the formamido derivatives, but the effect is larger for the dissociation kinetics. The formamido group thus strongly affects the interaction of polyamides with DNA and changes the preferred DNA sequences that are recognized by a specific polyamide stacked dimer.

Base Sequence↗
PubMed2026

A noncontiguous code for RNA-guided DNA recognition at the origin of CRISPR-Cas.

CRISPR-Cas provides RNA-mediated adaptive immunity, but how its first RNA-guided effector arose is unclear. In this study, we report the discovery of Viral Interference Programmable Repeat (VIPR) systems consisting of a Vipr protein ancestral to the earliest CRISPR-Cas effectors and VIPR RNAs (vrRNAs) comprising alternating GGY/NN motifs. Unlike canonical guide RNAs that pair with target nucleic acids through contiguous complementarity, vrRNAs recognize double-stranded DNA through a noncontiguous code in which the variable NN dinucleotides collectively specify a gapped target sequence. Natural vrRNA targets suggest that VIPR systems act against competing phages, and we demonstrate programmable phage defense by redirecting the complex for transcriptional repression. These results suggest that adaptive immunity originated from ancient warfare between viruses, revealing a previously unidentified logic for encoding information in sequence.

CRISPR-Cas Systems↗
PubMed2003

Selective DNA recognition by the androgen receptor as a mechanism for hormone-specific regulation of gene expression.

The androgen receptor (AR) is a member of the highly conserved group of the class I steroid hormone receptors, a subgroup of the nuclear receptor superfamily of ligand-induced transcription factors. All class I receptors influence the expression of their target genes by binding three-nucleotide spaced partial palindromic repeats of the core 5'-TGTTCT-3' motif. The implication that all class I receptors activate transcription by binding similar DNA motifs, poses the problem of how the in vivo steroid-specificity of transcriptional control is achieved. The AR, however, is able to interact with DNA motifs that are divergent from the classical hormone response elements. We will describe this AR-specific DNA interaction in the context of the general mechanisms that dictate the sequence-specificity of DNA-binding and dimerization of the nuclear receptors. The androgen receptor is the only steroid hormone receptor that is able to interact with response elements that are essentially arranged as a direct repeat of the 5'-TGTTCT-3' monomer binding element. We propose that the DNA-binding domain of the AR can interact with these androgen-specific response elements in a head-to-tail conformation, similar to many other nuclear hormone receptors. The fact that subtle differences in the sequence of response elements can dictate androgen-specific responses is a new and intriguing finding. It creates new possibilities in the research on hormone-selective action and provides a new angle in the search for selective ligands or co-factors that might influence androgen receptor action via either type of DNA motif.

Amino Acid Sequence↗
PubMed1999

Immunoglobulin motif DNA recognition and heterodimerization of the PEBP2/CBF Runt domain.

The polyomavirus enhancer binding protein 2 (PEBP2) or core binding factor (CBF) is a heterodimeric enhancer binding protein that is associated with genetic regulation of hematopoiesis and osteogenesis. Aberrant forms of PEBP2/CBF are implicated in the cause of the acute human leukemias and in a disorder of bone development known as cleidocranial dysplasia. The common denominator in the natural and mutant forms of this protein is a highly conserved domain of PEBP2/CBF alpha, termed the Runt domain (RD), which is responsible for both DNA binding and heterodimerization with the beta subunit of PEBP2/CBF. The three-dimensional structure of the RD bound to DNA has been determined to be an S-type immunoglobulin fold, establishing a structural relationship between the RD and the core DNA binding domains of NF-kappaB, NFAT1, p53 and the STAT proteins. NMR spectroscopy of a 43.6 kD RD-beta-DNA ternary complex identified the surface of the RD in contact with the beta subunit, suggesting a mechanism for the enhancement of RD DNA binding by beta. Analysis of leukemogenic mutants within the RD provides molecular insights into the role of this factor in leukemogenesis and cleidocranial dysplasia.

Cleidocranial Dysplasia↗
PubMed1992

The yeast alpha 1 and MCM1 proteins bind a single strand of their duplex DNA recognition site.

The yeast cell type regulator alpha 1 cooperates with a constitutive factor, MCM1 protein, to recognize the promoter and activate transcription of several alpha-specific genes. I show here that the alpha 1 and MCM1 proteins bind specifically to one of the two strands of their recognition sequence. This single-strand-binding activity shares several characteristics with the duplex-binding properties of these proteins: (i) the MCM1 protein binds alone to single-stranded and duplex sequences of both the alpha-specific (P'Q) and a-specific (P) binding sites; (ii) the alpha 1 protein requires both the MCM1 protein and the Q sequence to bind either single-stranded or duplex DNA; (iii) the alpha 1 protein stimulates binding of the MCM1 protein to both single-stranded and duplex DNAs; and (iv) the affinities of the proteins for single-stranded and duplex DNAs are comparable.

Base Sequence↗
PubMed1993

The function of individual zinc fingers in sequence-specific DNA recognition by transcription factor IIIA.

Mutations in Zn(2+)-coordinating histidine residues have been used to generate forms of transcription factor IIIA containing structural disruptions in single zinc fingers. These mutant proteins have been analyzed with respect to the structural and functional independence of individual zinc fingers in TFIIIA. They have also been used to assess the energetic contributions to binding and the sites of interaction of each of the nine zinc fingers of TFIIIA with the 5 S rRNA gene. The results are surprising and suggest a complex mode of binding in which the interactions of structurally independent zinc fingers with the DNA substrate are non-uniform and functionally interdependent in a way that may help to explain some of the unusual properties of TFIIIA.

Animals↗
PubMed2006

Neomycin-capped aromatic platforms: quadruplex DNA recognition and telomerase inhibition.

A series of aminoglycoside-capped macrocyclic structures has been prepared using intramolecular bis-tethering of neomycin on three aromatic platforms (phenanthroline, acridine, quinacridine). Based on NMR and calculations studies, it was found that the cyclic compounds adopt a highly flexible structure without conformational restriction of the aminoglycoside moiety. FRET-melting stabilization measurements showed that the series displays moderate to high affinity for the G4-conformation of human telomeric repeats, this effect being correlated with the size of the aromatic moiety. In addition, a FRET competition assay evidenced the poor binding ability of all macrocycles for duplex DNA and a clear binding preference for loop-containing intramolecular G4 structures compared to tetramolecular parallel G4 DNA. Finally, TRAP experiments demonstrated that the best G4-binder (quinacridine ) is also a potent and selective telomerase inhibitor with an IC(50) in the submicromolar range (200 nM).

Acridines↗
PubMed1994

Synthetic alpha-helical peptides incorporating intercalators for DNA recognition.

The design and synthesis of a water-soluble 14-residue peptide, in which a quinoline intercalator is attached to the peptide backbone via alkylation of a central cysteine residue, is reported. 600 MHz 1H NMR spectroscopy and circular dichroism indicate that the peptide forms a nascent helix in aqueous solution, ie. an ensemble of turn-like structures over several adjacent residues in the peptide. A large number of sequential dNN(i, i+1) connectivities were observed in NOESY spectra, and titration of trifluoroethanol into a solution of the peptide resulted in the characteristic CD spectrum expected for an alpha-helix. At low DNA concentrations, CD spectroscopy indicates that this helical conformation is stabilized, presumably due to folding of the peptide in the major groove of DNA.

Amino Acid Sequence↗
PubMed2002

Specificity in DNA recognition by phage integrases.

The lambda-related (lambdoid) coliphages are related to one another by frequent natural recombination and maintain a high level of functional polymorphism for several activities of the phages. Arguments are presented that the polymorphism of the integration module results from selection (presumably frequency-dependent) for new (not improved) specificities of site recognition. Analysis of phages lambda and HK022 by Weisberg and collaborators previously showed that changes in five noncontiguous amino acids could switch site recognition specificity. Phage 21 and defective element e14, which integrate at the same site, differ in recognition specificity for both core and arm sites. In vitro assays of e14 and 21 insertion and excision confirm this conclusion. Inhibition by ds arm site oligonucleotides defines the sequence specificity more precisely.

Bacteriophage lambda↗