Gas-Phase reactivity of Pb2+ ions with adenine, thymine and uracil

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Gas-Phase reactivity of Pb2+ ions with adenine, thymine and uracil

Jean-Yves Salpin, Sébastien Guillaumont, Jeanine Tortajada

To cite this version:

Jean-Yves Salpin, Sébastien Guillaumont, Jeanine Tortajada. Gas-Phase reactivity of Pb2+ ions with adenine, thymine and uracil. 16th International Mass Spectrometry Conference, 2003, Edimbourg, United Kingdom. 2003. �hal-00069112�



Methodology



Introduction

Gas-phase reactivity of Pb

2+

ions with adenine, thymine and uracil

Laboratoire Analyse et Environnement - UMR CNRS 8587



Results

Bâtiment des Sciences - Université d’Evry Val d’Essonne - Boulevard François Mitterrand - 91025 Evry Cedex - France

• Geometry optimization and vibrational analysis at B3LYP/6-31G(d,p) and G96LYP/6-31G(d,p) levels.

• NBO analysis.

• Many studies have shown that in physiological media, Pb(II) ions are determinant in the activity of some RNA-cleaving ribozymes . Moreover, it recently appeared that some ribozymes require the presence of two metal cations.(1,2) _{It was thus observed that in solution, Pb}2+ _{cations do not exhibit any particular reactivity with the DNA, whereas they induce the cleavage of the RNA in}

the presence of Mg2+_{. On the other hand, few is known about the intimate mechanisms associated with these interactions. In this context, we began a study in order to have a better}

understanding of these mechanisms, by considering in a first step the gas-phase reactivity between Pb2+ _{ions and three nucleobases: thymine (T, R=CH}

3), uracil (U R=H), and adenine (A).

• Electrospray/Triple quadrupole tandem mass spectrometer.

• Pb(NO₃)₂/nucleobase solutions (5.10-5 _mol.L-1_/10-4 _mol.L-1_{) in water or in a wather/methanol}

(50/50) mixture.



Mass Spectrometry

• Flow rate: 5l/min. • Temperature : 373 K.

Jean-Yves Salpin, Sébastien Guillaumont, Jeanine Tortajada



DFT calculations

• Use of the Stuttgart quasi-relativistic pseudo-potential to describe the lead atom.

HN N H O O R 4 6 1 2 3 5 _N N N N H NH₂ 1 2 3 5 4 6 7 8 9 10

• MS/MS experiments : collision gas N₂ (P  2 10-5 _{Torr ).}

• Formation of [Pb(B)_n – H]+ _{ions (n=1-3), the most intense ion corresponding to the complexation of a single nucleobase. • Characterization of ions [Pb(B) - H]}+ _{: MS/MS experiments and theoretical calculations.}

• Results obtained for thymine and uracile are totally comparable.



Pyrimidic bases : thymine and uracil

• MS/MS spectrum of the ion [Pb(T)-H]+ _{(m/z 333)}

m/z 225 [PbOH]+ [Pb(T)-H]+ [Pb(NCO)]+ -NCO. - Pb m/z 333 m/z 290 m/z 263 m/z 82 m/z 250 m/z 208 m/z 209 PbH+ (m/z 225) (m/z 334) (m/z 290) (m/z 263) (m/z 82) (m/z 251) Pb+ (m/z 208) (m/z 209) -C₅H₄N₂O -HNCO -C₄H₅NO -HCN -C₄H₃NO

• The elimination of HNCO specifically induces the carbon C(2).

• The ion [PbNCO]+ _{specifically induces the carbon}

N°2.



Puric base : adenine

• 12 structures considered. Deprotonation of the least acidic site (N₃) corresponds to the most stable structures. Monodentate species are not favored.

0

+14.6

+16.9

+25.1

+24.4

+28.8

_+30.5

• Tautomeric forms are particularly stable.

0

+12.1

+12.8

+24.9

+25.5

+69.5

+66.7

• Relative energies refined with an extended 6-311+G(3df,2p) like basis set (given below in kJ.mol-1 _).

• [Pb(T)-H]+ _{ions correspond to a mixture of several}

structures.

 DFT calculations

• B3LYP and G96LYP give similar relative energies.

• Study of modified pyrimidic bases (MB): MS/SM spectrum of [Pb(MB)-H]+

50 100 150 200 250 300 m/z, amu 0% 10% 20% 30% 40% 50% 60% 70% 80% 90% 100% Re l. Int. (%) 250 290 333 208 82 249 HN N O O CH3 50 100 150 200 250 300 350 0% 20% 40% 60% 80% 100% Re l. Int. (%) 95 335 266 241 208 ₂₇₆ 68 HN N H O S m/z, amu m/z, amu 50 100 150 200 250 300 0% 10% 20% 30% 40% 50% 60% 70% 80% 90% 100% Re l. Int. (%) 250 333 290 249 208 225 HN N H O O CH₃ 0% 20% 40% 60% 80% 100% Re l. Int. (%) 50 100 150 200 250 300 350 m/z, amu 127 335 275 250 208 292 95 265 241 HN N H S O 100 150 200 250 300 m/z, amu 0% 20% 40% 60% 80% 100% Re l. In t. ( % ) 251 334 263 208 ₂₉₀ 225 82 2-13_C-thymine HN N H O O CH₃ * 100 150 200 250 300 m/z, amu 0% 20% 40% 60% 80% 100% Re l. In t. ( % ) 250 333 263 208 ₂₉₀ 225 82 thymine HN N H O O CH₃

• Clear-cut distinction of methyl-uracil isomers. These MS/MS spectra also provide useful information about the mechanisms associated with the diferent fragmentations.

• Unambiguous characterization of 2-thio and 4thio-uracil.

• NBO analysis : net charge of Pb 1.52-1.53.

natural electronic configuration of Pb 6s(1.93)6p(0.53)sp0.27

Pb(II) lone pair : 6p contribution of 3.3 to 3.6 % characteristic of hemi-directed structures. (3)

T1

T2

T3

T4t

N₃ C₂ N₁ H C₆ C₅ C O O CH₃ H Pb N₃ C₂ N₁ H C₆ C₅ C₄ O O CH₃ H Pb N₃ C₂ Pb O + m/z 333 m/z 250 + + + N₁ C₆ C₅ C₄ O CH₃ H H N₃ C₂ N₁ C₆ C₅ C₄ O O CH₃ H H Pb

+

N₃ C₂ N₁ C₆ C₅ C₄ O O CH₃ H H Pb

+

+

m/z 333

m/z 290

T2

T3

 Possible mechanisms of dissociation

• MS/MS spectrum of the ion [Pb(A)-H]+ _{(m/z 342)}

• The fragmentations leading to the reduction of lead are particularly favorable (m/z 134 and 208).

100 150 200 250 300 350 m/z, amu 0% 20% 40% 60% 80% 100% Re l. In t. ( % ) 342 208 315 134 288 N N N N H H₂N adenine N N N N H H₂N * 100 150 200 250 300 350 m/z, amu 0% 20% 40% 60% 80% 100% Re l. In t. ( % ) 343 208 315 135 288 316 2-13_C-adenine N N N N N H H H H Pb N C N N N N H H H H Pb N C H + m/z 342 * * m/z 315 N N N N N H H H H * - Pb m/z 134 * N N N N H H H Pb  DFT calculations

 Possible mechanisms of dissociation

[C₅H₄N₅]+ m/z 342 (m/z 343) m/z 288 m/z 134 (m/z 135) m/z 208 [Pb(A)-H]+ m/z 315-316 Pb+ - HCN (85%)

*

- HCN (15%)

*

-Pb -C₅H₄N₅

(1) T. Pan, O. C. Uhlenbeck, Nature 358 (1992) 560.

(2) M. H. Kim, M. Katahira, T. Sugiyama, S. Uesugi J. Biochem. 122, 1062 (1997).



References

(3) L. Shimony-Livny, J. P. Glusker, C. Bock Inorg. Chem. 37

(1998) 853.

A1

A2

A3

A5t

• 24 distincts minima (monodentates or bidentates) were located. The results are comparable to those obtained for thymine or uracil. Removal of the most acidic proton (N9) leads to a complex 70 kJ.mol-1 _{less stable than the}

tautomeric form A5t. The local charge ranges between 1.39 for A1 and 1.51 for A3. The electron transfer is particularly important when lead binds to the exocyclic nitrogen (N10). Like for pyrimidic bases, the natural electron configuration of the metal is characteristic of hemi-directed structures.