Proton exchange of nucleic acids: The amino protons of 5′-AMP and poly A

McConnell, Bruce; Raszka, Matthew; Mandel, M.

doi:10.1016/0006-291x(72)90547-5

Cited by 9 publications

(6 citation statements)

References 9 publications

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“…Adenosine amino proton resonances are usually broadened by intermediate chemical exchange through rotation about the C–N bond [71,72,74]. However, for the A23–U12 pair we observe two distinct resonances for the A23 NH2 resonances, as a well as a perturbed, downfield 15 N chemical shift.…”

Section: Discussionmentioning

confidence: 69%

“…5A). Adenine amino resonances are normally broadened by chemical exchange about the C-N bond, whereas cytosine amino protons in Watson-Crick pairs usually give distinct resonances for hydrogen bonded (downfield) and non-hydrogen bonded protons [67][68][69][70][71][72][73][74][75]. The 15 N-1 H HSQC, focused on amino 15 N resonances, reveals the presence of novel shifted residues that mark the tertiary structure of tRNA.…”

Section: Resultsmentioning

confidence: 99%

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Probing the conformation of human tRNA₃^Lys in solution by NMR

Puglisi

2007

FEBS Letters

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Human tRNA Lys 3 acts as a primer for the reverse transcription of human immunodeficiency virus genomic RNA. To form an initiation complex with genomic RNA, tRNA Lys 3 must reorganize its secondary structure. To provide a starting point for mechanistic studies of the formation of the initiation complex, we here present solution NMR investigations of human tRNA Lys 3 . We use a straightforward set of NMR experiments to show that tRNA Lys 3 adopts a standard transfer ribonucleic acid tertiary structure in solution, and that Mg 2+ is required for this folding. The results underscore the power of NMR to reveal rapidly the conformation of RNAs.

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Section: Discussionmentioning

confidence: 69%

Section: Resultsmentioning

confidence: 99%

Probing the conformation of human tRNA₃^Lys in solution by NMR

Puglisi

2007

FEBS Letters

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“…The half-height widths of the 300-MHz amino proton signals were somewhat less than those of corresponding experiments at 100 MHz (McConnell et al, 1972). However, we consider the line widths to be the same for the two magnetic fields, since the 5'-AMP samples used for the 300-MHz spectra were highly purified and were measured in a 2-mm capillary, which eliminates the water spinning side bands that contribute to the overall line width in the 100-MHz experiments.…”

Section: Pmr Experimental Proceduresmentioning

confidence: 61%

“…This problem was not encountered with the cyclic compounds and with 3 '-AMP, whose signal to noise ratio for the amino protons are several times larger and the half-height widths are about 10 Hz. Assignment of this signal to the amino protons (with 5'-AMP) was verified with the use of the 300-MHz spectrometer,2 which provided sufficient separation of the amino and water proton peaks (550 Hz) for accurate comparison of the signal area (2 protons) to that of the H-8 and H-2 single protons (3.7 and 3.3 ppm, respectively) (McConnell et al, 1972). The similar assignment for the amino proton signal of 5'-GMP relative to H-8 and H-l' is in agreement with the tentative assignment made by Miles et al (1963) for this compound in aqueous solution.…”

Section: Pmr Experimental Proceduresmentioning

confidence: 94%

Proton exchange of nucleic acids. Amino protons of mononucleotides

McConnell¹,

Seawell²

1972

Biochemistry

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Exchange of the amino protons of 2':3'-cAMP, 2' :3'-cGMP, 5'-AMP, 5'-GMP, 3'-AMP, and adenosine can be measured in aqueous solution with the aid of the 100-MHz nmr spectrometer. The variation in line width of the -NH2 proton resonance with p H is used to estimate empirically the second-order rate constants for proton-exchange catalysis by H30+ and OH-, which are in the range 106-108 M-' sec-l. These estimates correspond to rates for intrinsic exchange that are several orders of magnitude less than rates for aliphatic T h e ways in which the exchangeable purine and pyrimidine protons of helical DNA can be hidden from contact solvent by secondary structure appear to be few and well defined. A space-filling model of the double helix would show one proton of each of the amino pairs sterically accessible to solvent in the helical grooves and free from participation in intramolecular hydrogen bonding. In contrast, the second amino proton and the imino protons of the guanine and thymine rings are much less accessible to solvent molecules because of their location and their involvement in hydrogen bonds between the complementary strands. Therefore, initial observations that exchange of the hydrogens of helical polynucleotides was slow and accessible to measurement by gel filtration techniques is significant for the use of hydrogen exchange as a general probe of macromolecular structure (Printz andvon Hippel, 1965, 1968;Englander and Englander, 1965). The DNA helix might represent a relatively simple model for obtaining information, which could be applied not only to questions of its own structure, but to interpretation of HX data of the more complex proteins (Hvidt and Nielsen. 1966). Potentially, this information is of two kinds: (1) the ciniount of macromolecular structure, which is obtained by counting "slow" hydrogens and (2) the motility of the structure, which is derived from exchange rates (McConnell and von Hippel, 1971). Both types of information require a quantitative knowledge of the hydrogen-exchange rate in the absence of structure, which has not been available for nucleic acids.Consideration of the general aspects of DNA and polynucleotide hydrogen-exchange data that has been gathered to date indicates that there is a need for measurements of the intrinsic exchange of the purine and pyrimidine protons before a mechanism can be elucidated for exchange in the macromolecule, especially for the purpose of describing the nature and position of the structure motility in the sequence of events that initiate exchange. A comprehensive list of these general features would include the following. (1) Structural and chemand aromatic amines, and account largely for the fact that "outside" amino protons of helical polynucleotides can be measured by slow kinetic techniques. The pH-exchange profiles do not coincide for the adenine and guanine -NH2 protons, which accounts for the different kinetic classes of exchangeable hydrogen from the DNA helix. Amino proton exchange is catalyzed by phosphate and imidazole and is highly ...

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“…In fact, their intrinsic exchange rates are several orders of magnitude lower than the diffusion-controlled rates for aliphatic and aromatic amines. One can find appropriate conditions (nucleotide concentration, temperature and pH), which vary with the nucleotide, at which the exchange rate is at the slow exchange limit [17]. This is given experimentally by the independence of the line width for the amino protons resonance on the peak separation between -NHz and H2O protons.…”

Section: Sh$t Data In H20mentioning

confidence: 99%

Nuclear-Magnetic-Resonance Studies of 5'-Ribonucleotide and 5'-Deoxyribonucleotide Conformations in Solution Using the Lanthanide Probe Method

et al. 1978

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icleo tide WILLIAMSThe conformations of the metal-bound 5 '-ribonucleotides and 5 '-deoxyribonucleotides in aqueous solution at different pH values have been studied using the lanthanide probe method. The conformational analysis, based on mixing different conformations in fast exchange within the nuclear magnetic resonance time scale, agrees well with the results from coupling constants, nuclear Overhauser effects and spin-lattice relaxation times, obtained for the metal-fixed systems. The equilibrium between the two basic conformational combinations for the 5 '-nucleotides, anti-(N G? S ) -gg-g'g' and syn-(NG?S)-gt-g'g' depends on the nature of the furanose ring, the base and also on the state of base protonation and phosphate ionization. The effect of base protonation is particularly strong for the guanine nucleotides.The conformations of free 5 '-ribonucleotides and deoxyribonucleotides in solution have been extensively studied by nuclear magnetic resonance (NMR) spectroscopy, due to the importance of these molecules as the simplest constituents of naturally occurring nucleic acids. Most of these studies have been based on the interpretation of experimental NMR parameters for the isolated nucleotides, such as chemical shifts, vicinal spin-spin coupling constants, nuclear Overhauser enhancements and nuclear relaxation rates [I, 11 -13,281 (and references in [I]). A different general method for the determination of the conformation of metal-bound flexible molecules in solution, using the lanthanide cations (Ln3 ') as shift and relaxation NMR probes has been described and applied to some nucleotide monophosphatates [2-41.In this paper the lanthanide probe method is applied to the comparative study of the conformations in aqueous solution of the eight metal-bound 5'-mOnOnucleotides which occur in ribonucleic and deoxyribonucleic acids : the nucleotides of adenosine (5'-AMP), guanosine (5'-GMP), cytidine (5'-CMP), uridine (5 '-UMP), 2'-deoxyadenosine (5 '-dAMP), 2'-deoxyguanosine (5'-dGMP), 2'-deoxycytidine (5'-dCMP) and thymidine (5'-dTMP).The conformational conclusions obtained from the lanthanide data are compared with independent NMR Ahbrevintion. NMR, nuclear magnetic resonance work, especially coupling constants and nuclear Overhauser enhancement data, as there must be at least a qualitative agreement for the metal-bound systems between the conclusions from the different methods [5,6]. From this comparison, an assessment of the different NMR conformational methods in solution is attempted. Finally the results in aqueous solution are compared with the available information on the crystal structure of metal-bound 5 '-mononucleotides from X-ray diffraction data and with the data on the metal-free systems. MATERIALS AND METHODS'H NMR spectra were obtained at 90 MHz using a Bruker HFX 90 spectrometer and at 270 MHz using an Oxford Instruments superconducting magnet and a modified Bruker HFX 90 console. Both spectrometers were operated in the Fourier transform mode, using a Nicolet 1085 computer. The observation o...

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Proton exchange of nucleic acids: The amino protons of 5′-AMP and poly A

Cited by 9 publications

References 9 publications

Probing the conformation of human tRNA₃^Lys in solution by NMR

Probing the conformation of human tRNA₃^Lys in solution by NMR

Proton exchange of nucleic acids. Amino protons of mononucleotides

Nuclear-Magnetic-Resonance Studies of 5'-Ribonucleotide and 5'-Deoxyribonucleotide Conformations in Solution Using the Lanthanide Probe Method

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Proton exchange of nucleic acids: The amino protons of 5′-AMP and poly A

Cited by 9 publications

References 9 publications

Probing the conformation of human tRNA3Lys in solution by NMR

Probing the conformation of human tRNA3Lys in solution by NMR

Proton exchange of nucleic acids. Amino protons of mononucleotides

Nuclear-Magnetic-Resonance Studies of 5'-Ribonucleotide and 5'-Deoxyribonucleotide Conformations in Solution Using the Lanthanide Probe Method

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Probing the conformation of human tRNA₃^Lys in solution by NMR

Probing the conformation of human tRNA₃^Lys in solution by NMR