Dehydrogenation of nucleobases upon

electron attachment to isolated nucleotides

Steen Brøndsted Nielsen

Department of Physics and Astronomy

University of Aarhus

June 24-27, 2004, Lyon-France 

DNA DAMAGE

HOW DOES UV RADIATION CAUSE GENETIC DAMAGE ?

1) Direct absorption of light by the nucleobases

followed by physical and chemical reactions

and / or

2) Ionisation of water (H2O H2O+ + e) followed

by electron attachment to nucleobases

Modification of nucleobases may lead to loss of base-pairing specificity

Elucidate the processes at the single-molecule level.

Isolated damage to nucleotides, the basic building

blocks of DNA and RNA, upon electron attachment.

OUR APPROACH

TOOLBOX

ELECTROSPRAY IONISATION

MASS SPECTROMETRY:

ACCELERATOR INSTRUMENT

ION STORAGE RING ELISA

LASERS

QUANTUM CHEMICAL CALCULATIONS

BARE IONS

ELECTROSPRAY IONISATION

droplets

ELECTROSPRAY ION SOURCE

1 mbar 10-3 mbar 10-5 mbar 10-6 mbar

Heatedcapillary

ESI needle4kV

22-pole ion trap

Tube lensSkimmer

OctapoleLenses

Acceleration tube

GAS-PHASE EXPERIMENTS ON NUCLEOTIDE ANIONS

PROBLEM:

Attachment of an electron to a negative

ion is unlikely because of the repulsion

between the two negative charges.

SOLUTION:

High-energy collisions with sodium:

electron transfer from Na to the ion.

R 0

dianion

En

erg

y

Anion - electron distance R

dianion

Compare: the violent reaction of sodium with water !

Nielsen et al., J. Am. Chem. Soc. 125, 9592-9593 (2003).

Liu et al., Chem. Phys. Chem. 4, 1341-1344 (2003).

ACCELERATOR INSTRUMENT

ESIsource

Magnet

Na collision cell

Electrostaticanalyser

0 V

50 kV

Channeltrondetector

+

R = 2 mB = 1.4 T

T=500 K

COLLISION INTERACTION TIME: FEW FEMTOSECONDS

Na

-Nucleotide anion

105 m/s

1 nm

The electron transfer process is nearly vertical.

Electron attachment to AMP anions

O

OH OH

N N

NN

NH2

CH2O

O

P

O

OH

1

2

3

4

567

8

9

1'

2'3'

4'

5'

m/q 346

Ne

Na

Liu et al., Chem. Phys. Chem. 4, 1341-1344 (2003).

80 120 160 200 240 280 320 360

dianion

[AMPAH] [AMPO]

AMP

A

H2PO

4

PO3

AMP

[AMPAH]A

H2PO

4

PO3

m/q

164 166 168 170

High-energy collisions between dAMP anions and Na and Ne

100 150 200 250 300

m/q

dAMPdAMP2

fragment ionsm/q 330

Spectrum obtained after collisions between AMP(H2O)13 and Na

100 200 300 400 500 600

m/q

170 190 210 230 250

AMP(H2O)13

AMP2 AMP2(H2O)9

AMP

100 200 300 400 500 600

m/q

dG2

dT2

dC2

dA2

Electron transfer from Na to dinucleotide anions

sugar

sugar

phosphate

nucleobase

nucleobase

dianion

-

IS THE NUCLEOTIDE DIANION INTACT ?

H loss is observed upon electron attachment to nucleobases in vacuo.

C. Desfrancois, H. Abdoul-Carime, and J. P. Schermann, J. Chem. Phys. 104, 7792 (1996).S. Gohlke, H. Abdoul-Carime, and E. Illenberger, Chem. Phys. Lett. 380, 595 (2003). G. Hanel, B. Gstir, S. Denifl, P. Scheier, M. Probst, B. Farizon, M. Farizon, E. Illenberger, and T. D. Märk, Phys. Rev. Lett. 90, 188104 (2003). H. Abdoul-Carime, S. Gohlke, and E. Illenberger, Phys. Rev. Lett. 92, 168103 (2004).

H loss is observed upon electron attachment to deoxyribose in vacuo.

S. Ptasińska, S. Denifl, P. Scheier, and T. D. Märk, J. Chem. Phys. 18, 8505 (2004).

1.5 1 0.5 0m/q

# of D in AMP

0

1

2

3

Intact dianion H D

AMP dissolved in CD3OD:

Selection of deuterium-

labelled ions for collision

experiments

O

OH OH

N N

NN

NH2

CH2O

O

P

O

OH

1

2

3

4

567

8

9

1'2'3'

4'

5'

Exchangeable hydrogens

Liu et al., J. Chem. Phys., issued for Sept. 1 (2004).

WHAT IS THE ORIGIN OF THE HYDROGEN ?

POH phosphoric acid group

OH sugar

NH2 nucleobase

CH sugar or nucleobase

Electron attachment to nucleotides:

dehydrogenation at nitrogen sites

N

NH

O

OCH3

N

NH

O

O

Thymine Uracil Adenine Cytosine Guanine

N N

N N

NH2

N

N

O

NH2

N N

N NH

O

NH2

1 2 3

0 1 2 30.0

0.1

0.2

0.3

0.4

(Å

2 )

Number of nitrogen hydrogens

RNA nucleotide DNA nucleotide

Cross section for formation of dehydrogenated dianion

• H-loss from N

• No D-loss from C

H. Abdoul-Carime, S. Gohlke, and E. Illenberger,

Phys. Rev. Lett. 92, 168103 (2004).

Dissociative electron attachment to deuterated thymine

THE DNA DOUBLE HELIX

Watson-Crick base pairs Base mispairing

Biological relevance?

WHAT IS THE LIFETIME OF THE DIANION ?

1 m

Micro-channel platedetector

Accelerator with electrospray ion source

Magnet

Ion bunch

Injection

ELectrostatic Ion Storage Ring Aarhus (ELISA)

S.P. Møller, NIM A 394, 281 (1997).

J.U. Andersen, J.S. Forster, P. Hvelplund, T.J.D. Jørgensen, S.P Møller, S. Brøndsted Nielsen, U.V. Pedersen, S. Tomita, and H. Wahlgreen, Rev. Sci. Instrum. 73, 1284-1287 (2002).

Channeltron

Sodium

0.0 0.5 1.0 1.5 2.0

100

101

102 298 K 203 K

Dec

ay r

ate

[a.u

.]

Time [ms]

= 0.2 ms

Electron autodetachment from the AMP dianion

Liu et al., J. Chem. Phys., issued for Sept. 1 (2004).

1 m

Micro-channel platedetector

Accelerator with electrospray ion source

Magnet

Ion bunch

Injection

Channeltron

Long-lived component ?

Dump of a remaining beam in the micro-channel plate detector after 34 ms of storage time.

Sodium

34.00 34.05 34.10 34.150

30

60

90

Cou

nts

Time [ms]

40 s

Dump of beam in the detector after 34 ms of storage time

AMP2

(m/q 172.5)

13C-AMP2

(m/q 173)

CONCLUSIONS

• Electron transfer to nucleotide anions occurs in

collisions with sodium vapour.

• Electron attachment leads to dehydrogenation of

the nucleobase nitrogens.

• The dehydrogenated dianion is longlived.

Other fragmentation channels than H-loss upon electron attachment ?

ACKNOWLEDGEMENTS

University of Aarhus:

Preben Hvelplund Jens Ulrik Andersen

Shigeo Tomita Bo Liu

Jimmy Rangama

Leopold-Franzens Universität Innsbruck:

Paul Scheier Gabriel Hasan

FUNDING

The Danish Natural Science Research Council

Aarhus Center for Atomic Physics (ACAP)