Resonance assignments of non-exchangeable protons in B type DNA oligomers, an overview

The solution structure of the deoxydecanucleotide [d(GAGTUACGA)] . [d(GAGTUACGA)] has been determined by NMR methods. This duplex, which contains six G . A mismatches and four Watson-Crick base pairs, is thermodynamically more stable than a decamer where T . A base pairs are substituted for the G . A mismatches, and is less stable than the duplex that contains G . C base pairs. Circular-dichroism spectroscopy indicates an overall B-like conformation for the decamer, but stronger than usual base staclung. 'H-NMR spectroscopy revealed that the N1H groups of the mismatched guanine residues are not hydrogen bonded, and 31P-NMR showed the presence of B,, phosphate conformations for the GpA steps. Detailed analysis of the NMR data showed that all nucleotides have anti glycosidic torsion angles and S type sugar puckers. The G .

Section: Solution Structure Of [D(gagt=acga)] -[D(gagtgaacm)]mentioning

confidence: 79%

Section: Solution Structure Of [D(gagt=acga)] -[D(gagtgaacm)]mentioning

confidence: 94%

Section: Conformationmentioning

confidence: 99%

See 1 more Smart Citation

Properties of multiple G · A mismatches in stable oligonucleotide duplexes

Lane¹,

Ebel²,

Brown³

1994

“…1). All non-exchangeable protons except the H5' and H5" protons were assigned in the four oligonucleotides (Table 1) (Van de Ven and Hilbers, 1988). Upon methylation, both (CA) and (CT) present a high field shift of 0.15 ppm for their C4 H6 proton related to the inductive effect of the methyl group within the pyrimidine ring.…”

Section: Analysis Of Non-exchangeable Protonsmentioning

confidence: 99%

Sequence Dependent Effects of CpG Cytosine Methylation

Lefebvre

Mauffret

Antri

et al. 1995

The impact of cytosine methylation in the central CpG step of two closely related octanucleotide duplexes d(CATCGATG), and d(CTTCGAAG), was examined by 'H-NMR and 31P-NMR experiments, and a quantitative structural analysis was performed using the NOE-derived distances, the sugar puckers and the E torsion angles. The two starting oligonucleotides displayed a B-DNA conformation with, however, significant local structure differences. Although the methylated oligonucleotides retained their B-DNA conformation, different structural and thermal stability effects were observed. The magnitude of the methylation effects was to depend on the initial conformation of the CpG site, which is governed by the nature of the dinucleotide AT or TT located on the CpG flanks. As an example of sequence dependence, the methylation of CpG entailed larger conformational variation in d(CATCGATG), than in d(CTTCGAAG),. In this study, the 'H and 31P chemical-shift parameters averred as extremely sensitive probes for detecting subtle conformational changes. Finally, our comparative results may aid our understanding of the structural and related biological effects produced by cytosine methylation in DNA.

“…The major cause of chemical shift changes in the non-exchangeable protons of the DNA is probably changes in the orientation of the base pairs and their immediate neighbours, and hence changes in the ring current effects [34). For the exchangeable protons, changes in exchange rates with solvent, or in hydrogen bonding may also occur giving large changes in shift.…”

Section: Discussionmentioning

confidence: 99%

NMR Studies of the Escherichia coli Trp Repressor ·trpR^s Operator Complex

Evans

Jaseja

Jeeves

et al. 1996

To understand the specificity of the Escherichia coli Trp repressor for its operators, we have begun to study complexes of the protein with alternative DNA sequences, using 'H-NMR spectroscopy. We report here the 'H-NMR chemical shifts of a 20-bp oligodeoxynucleotide containing the sequence of a symmetrised form of the trpR operator in the presence and absence of the holorepressor. Deuterated protein was used to assign the spectrum of the oligodeoxynucleotide in a 37-kDa complex with the Trp holorepressor. Many of the resonances of the DNA shift on binding to the protein, which suggests changes in conformation throughout the sequence. The largest changes in shifts for the aromatic protons in the major groove are for A15 and G16, which are thought to hydrogen bond to the protein, possibly via water molecules. We have also examined the effect of DNA binding on the corepressor, tryptophan, in this complex. The indole proton resonance of the tryptophan undergoes a downfield shift of 1.2 ppm upon binding of DNA. This large shift is consistent with hydrogen bonding of the tryptophan to the phosphate backbone of the trpR operator DNA, as in the crystal structure of the holoprotein with the trp operator.Keywords: Trp repressor; deuteration ; trp operator.The Escherichia coli Trp repressor is activated by tryptophan to bind to at least five operators in the E. coli genome, namely the trpEDCBA (trpO), aroH, aroL, trpR and mtr operators (reviewed in [I]) [ 2 ] . It thereby represses initiation of transcription of genes involved in tryptophan uptake and biosynthesis in response to intracellular levels of tryptophan. As one of the smallest proteins whose binding to a specific DNA sequence is allosterically controlled, the Trp repressor has been studied extensively by a variety of genetic and physical methods; however, the basis for its DNA selectivity remains controversial.The structures of the protein and several of its complexes have been determined by X-ray crystallography [3-71 and, to lower resolution, by 'H and heteronuclear NMR spectroscopy [8-131. The protein belongs to the helix-turn-helix family of dimeric DNA-binding proteins. Tryptophan binds between the core of the molecule and the DNA-binding helices in each subunit, changing their orientation so that they are better positioned to bind to the DNA. In the crystal structure of the holoprotein bound to a double-stranded oligodeoxynucleotide containing a symmetrised version of the trpO operator sequence, the corepressor tryptophan also interacts directly with the phosphate backbone of the DNA [6]. Surprisingly in this structure there are no direct hydrogen bonds or van der Waals' interactions between the protein and the base pairs of the DNA known to be important for sequence-specific binding. There are several water-mediated interactions that could be important for specificity. Alternatively, specificity could arise from the interactions of the protein with the phosphate backbone ('indirect readout'). This result has been controversial: an NMR study of the same co...