Sequential resonance assignments in proton NMR spectra of oligonucleotides by two-dimensional NMR spectroscopy

Scheek, Ruud M.; Boelens, Rolf; Russo, Nino; Boom, Jacques H. van; Kaptein, Robert

doi:10.1021/bi00302a006

Cited by 267 publications

(186 citation statements)

References 14 publications

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“…These assignments were complete with the exception of some ambiguity in distinguishing between some of the H 2' and H 2" pairs and some of the H 5' and H5" pairs in a given mononucleotide. As discussed by others [4,9,13,211, in a B-type nucleic acid helix the internuclear separation between a base proton and its own H2' (i.e. on the sugar ring in the same mononucleotide residue as the aromatic base) is less than that of the base proton to its own H 2".…”

Section: Assignmentsmentioning

confidence: 90%

Investigation of the solution structure of a DNA octamer [d(GGAATTCC)]₂ using two‐dimensional nuclear Overhauser enhancement spectroscopy

Broido

James

Zon

et al. 1985

European Journal of Biochemistry

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Proton two-dimensional nuclear Overhauser enhancement (2 D NOE) spectra in the pure absorption phase were obtained at 500 MHz for [d(GGAATTCC)], in aqueous solution at a series of mixing times. The experimental data were analyzed by comparison with theoretical spectra calculated using the complete 70 x 70 relaxation matrix including all proton dipole-dipole interactions and spin diffusion [Keepers, J. W. &James, T. L. (1984) J . M a p . Reson. 57,404-4261. The theoretical spectra at each mixing time were calculated using two structures: a standard B-form DNA structure and an energy-minimized structure based on the similarity of the six internal residues of the title octamer with those of the dodecamer [d(CGCGAATTCGCG)],, for which the crystal structure has been determined. Neither the standard B-form nor the energy-minimized structure will yield theoretical 2 D NOE spectra which accurately reproduce all peak intensities in the experimental spectra. However, many features of the experimental spectra can be represented by both the B-form and the energy-minimized structure. Sequencedependent structural characteristics are manifest in the 2 D NOE spectra, in particular at the purine-pyrimidine junction as noted previously in the crystal structure. On the whole, the energy-minimized structure appears to yield theoretical 2 D NOE spectra which mimic many, if not all, aspects of the experimental spectra. All 2 D NOE data were consistent with nanosecond correction times as implied by proton spin-lattice relaxation time measurements. But better fits of some of the 2 D NOE data using small variations in an effective isotropic correlation time suggest that there may be some local variations in mobility within the octamer duplex structure in solution.With recent advances in DNA synthesis enabling the preparation of DNA oligomers of known, specific sequence [l -41, X-ray crystallographers are examining DNA oligomers of different sequences and reporting details of sequence-dependent crystalline conformations [3, 5 -81. Questions regarding the structure of these DNA fragments in solution are also being addressed. These questions focus on the extent of preservation of the crystal structures in solution and on the manner in which the solution and crystal structures differ. On a more general level, discussion also focusses on whether or not it is accurate to speak of a molecular structure in solution and by what means may such a structure be determined. The continuing development of high-resolution NMR technology has both spurred such questions and been of great use in the quest for answers. Two-dimensional NMR (2D NMR), in particular, has developed to the point that proton spectra of biomolecules, spectra with perhaps hundreds of lines and once thought to be undecipherable, can now, in many cases, be totally assigned [I, 4,9 -111. Following assignment, both oneand two-dimensional NMR techniques enable spectroscopists to start probing in detail the solution conformations of these molecules.Correspondence to T. L. James, Departmen...

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Section: Assignmentsmentioning

confidence: 90%

Investigation of the solution structure of a DNA octamer [d(GGAATTCC)]₂ using two‐dimensional nuclear Overhauser enhancement spectroscopy

Broido

James

Zon

et al. 1985

European Journal of Biochemistry

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“…It is seen that these are approximately the same in both forms, in accordance with known structural features (sugar S type, cytosine antiin both B and Z DNA). In contrast, the H-8/H-1' cross-peaks of the guanosines [residues (2), (4) and (6)] show a normal pattern in the B form, whereas extremely strong intraresidue H-8/H-1' cross-peaks are observed for the Z form, Fig.9A, left-hand side. This observation is again well explained by the X-ray model of Z DNA, which features syn guanosines with a concomitant short distance (approximately 0.24 nm) between H-8 and H-1'.…”

Section: Assignment Of the 111 Biz Mixturementioning

confidence: 95%

“…In fact, the corresponding cross-peaks allow a rapid mapping of all base and H-1' signals throughout the strand, Fig. 9 A right-hand side [4,7,25]. In the Z form such continuous mapping at first sight appears impossible.…”

Section: Assignment Of the 111 Biz Mixturementioning

confidence: 99%

The B and Z forms of the d(m⁵C‐G)₃ and d(br⁵C‐G)₃ hexamers in solution

Orbons¹,

Marel²,

Boom³

et al. 1986

European Journal of Biochemistry

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The non-exchangeable proton resonances of the hexadeoxynucleoside pentakisphosphates d(m5C-G)3 and d(b~-'c-G)~ in the B form as well as in the Z form were assigned by means of two-dimensional correlated spectroscopy and two-dimensional nuclear Overhauser enhancement spectroscopy. The complete proton NMR spectrum of the B form of the methylated compound was assigned in a pure 'H20 solution as well as in a 2H20/ C2H302H mixed solvent, containing 5 mM MgClz. In the latter solvent the B form occurs in slow equilibrium (on the NMR time scale) with the Z form, the resonances of which also were fully assigned. The proton resonances of the B and Z forms of the brominated fragment were assigned in a 2HzO/C2H302H solution containing 5 mM MgCI2. A new and general method is described for the sequential assignment of the non-exchangeable proton resonances of oligonucleotides in the Z form. A well-tested strategy is based upon a combined use of both techniques. In a first step COSY is used to identify groups of scalarly coupled nuclei (i. e. sugar protons belonging to a single residue). In the next step these sets of nuclei are assigned to a specific residue by means of nuclear Overhauser enhancements, observed between base and sugar protons. Although the NOESY spectrum alone usually contains sufficient information to complete the assignment without the aid of the COSY spectrum, the latter serves as an aid in avoiding pitfalls.Hitherto, no similar direct method has been available for the assignment of proton spectra of oligonucleotides in the Z form. Published assignments of the proton spectra of DNA fragments in the Z form thus far rely upon indirect techniques. In the present paper a new and general strategy based upon COSY and NOESY is proposed for the sequential assignment of oligonucleotides in the Z form. We will show that it is indeed possible to assign each of the 51 and the 48 non-equivalent resonances exhibited by the major species (B and Z) of the hexamers d ( m 5 C Q 3 and d(b~-%-G)~, respectively, to a specific proton. Both hexamers were chosen for this study because it is known that methylation or bromination of cytidine at the C-5 position facilitates the formation of the Z At this point it is well to remark that the polymorphism exhibited by duplexes of the d(C-G), . d(C-G), type necessitates a careful assignment and analysis of the NMR spectra of each stable molecular species. In the present paper we will loosely designate the low-salt form in aqueous solution as B DNA and the structure which becomes increasingly predominant when increasing amounts of C2H302H and/or MgC12 are added to the solution will be designated Z DNA.The conformational analysis of the respective B and Z forms will be reported in a companion paper [15]. There it will be shown that the interpretation of the NMR data indeed strongly supports these designations.form [12-141. MATERIALS AND METHODSThe hexamers d(m5C-G)3 and d(b~%-G)~ (Fig. 1) were synthesized via an improved phosphotriester method [16, 171. After purification the samples were ...

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“…As evident from 12.5 ppm ('4') also broadens with increase in temperature and disappears with presaturation; it is therefore assigned to 1NH of the terminal G1. Sequential resonance assignments of non-exchangeable protons were achieved through established procedures [15][16][17][18][19]. As an illustrative example of the resonance assignment procedure, Fig.…”

Section: Resultsmentioning

confidence: 99%

Evidence for A⁺(anti)–G(syn) mismatched base–pairing in d–GGTAAGCGTACC

Chary

Govil

et al. 1995

FEBS Letters

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Sequential resonance assignments in proton NMR spectra of oligonucleotides by two-dimensional NMR spectroscopy

Cited by 267 publications

References 14 publications

Investigation of the solution structure of a DNA octamer [d(GGAATTCC)]₂ using two‐dimensional nuclear Overhauser enhancement spectroscopy

Investigation of the solution structure of a DNA octamer [d(GGAATTCC)]₂ using two‐dimensional nuclear Overhauser enhancement spectroscopy

The B and Z forms of the d(m⁵C‐G)₃ and d(br⁵C‐G)₃ hexamers in solution

Evidence for A⁺(anti)–G(syn) mismatched base–pairing in d–GGTAAGCGTACC

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Sequential resonance assignments in proton NMR spectra of oligonucleotides by two-dimensional NMR spectroscopy

Cited by 267 publications

References 14 publications

Investigation of the solution structure of a DNA octamer [d(GGAATTCC)]2 using two‐dimensional nuclear Overhauser enhancement spectroscopy

Investigation of the solution structure of a DNA octamer [d(GGAATTCC)]2 using two‐dimensional nuclear Overhauser enhancement spectroscopy

The B and Z forms of the d(m5C‐G)3 and d(br5C‐G)3 hexamers in solution

Evidence for A+(anti)–G(syn) mismatched base–pairing in d–GGTAAGCGTACC

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Investigation of the solution structure of a DNA octamer [d(GGAATTCC)]₂ using two‐dimensional nuclear Overhauser enhancement spectroscopy

Investigation of the solution structure of a DNA octamer [d(GGAATTCC)]₂ using two‐dimensional nuclear Overhauser enhancement spectroscopy

The B and Z forms of the d(m⁵C‐G)₃ and d(br⁵C‐G)₃ hexamers in solution

Evidence for A⁺(anti)–G(syn) mismatched base–pairing in d–GGTAAGCGTACC