I. PEÑA, J.L. ALONSO
Grupo de Espectroscopia Molecular. Unidad asociada CSIC
Laboratorios de Espectroscopia y Bioespectroscopia
Edificio Quifima. Parque Científico
Universidad de Valladolid SPAIN

 Nucleosides bound to one or more phosphate groups
(nucleotides) are considered the molecular building blocks of DNA and RNA
 A nucleoside is composed of a nitrogen base bound to either ribose or deoxyribose via a beta-glycosidic linkage.
In particular, uridine is composed of uracil and ribose
 The jet-cooled rotational spectrum of uracil has been investigated using LA-MB-FTMW spectroscopy.
1 The diketo structure of uracil has been determined
H
O
H
O
H
N
3
O
H
H
N
O
H
N
H
N
O
N
1
H
DIKETO FORM
O
H
[1]J. Phys. Chem. A, 111, 3443 (2007)
N
O
N O
H H
PLAUSIBLE TAUTOMERS OF URACIL
N
HO
H
O
N
NH
O
URACIL
H
H
OH
O
H
OH
H
GLYCOSIDIC BOND
RIBOSE
URIDINE

 For D-ribose, eight different PYRANOSE FORMS have been recently characterized in gas phase using
CP-FTMW spectroscopy + Laser Ablation

H
O
N
H
NH
O
The most stable form of URACIL
H
OH
H
H
OH
O
H
OH
H
OH +
H
H
H
H
O
H
H
OH OH
OH OH
The most stable  and
β forms of RIBOSE
Why did nature choose the furanose and not the pyranose form of ribose in RNA????
O
NH
N
H
H
OH
H
H
OH
O
H
OH
H
O
O
NH
N O
HO
H
H
OH
O
H
OH
H
URIDINE
Furanose form of ribose in RNA

Some structural arguments could be based on the intramolecular interactions between the base and ribose moieties, which would stabilize the furanose form versus the pyranose form
The aim of the present work is the observation of the isolated nucleoside uridine in the gas phase, free from the bulk effects of the native envinroment, to reveal the biologically important uracil/ribose intramolecular interactions

A commercial sample of uridine (m.p.: 163-167ºC) was vaporized using the third harmonic of a 20 ps Nd:YAG laser. Products of the laser ablation were supersonically expanded in Ne and probed by broadband CP-FTMW
6-12GHz
120000 fids
6000 7000 8000 9000 10000 11000 12000
 / MHz
CP-FTMW + LA at GEM, Valladolid
S. Mata I. Peña, et al. J. Mol. Spectr. 280(2012) 91–96

PHOTOFRAGMENTATION PRODUCTS
2
Uracil
12

1
01
HC
3
N
7870 7875
 /MHz
7880 7885
Photofragmentation products such as uracil, glyceraldehyde, ethylene glycol, acetic acid, oxoacetic acid, formaldehyde, acrolein and cyanoacetylene seem to be dominant in the spectrum
6000 7000 8000 9000 10000 11000 12000
 / MHz
120000 fids
Is neutral uridine present in the supersonic expansion in an appreciable amount to be detected?

6000
Resultant spectrum without photofragmentation products
7000
10
19

9
18 transition
8000 rotational
6075
 /MHz
6080
9000
 / MHz
10000 11000
Assignment of
 a and
 b
-type R-branch progressions (with the quantum number J ranging from 4 to 17), as corresponding to one rotamer
12000
4
3
12
18
3
2
11
6
4
6
13
5
10
15
17
17
10
9
12
13
16
16
5
11
12
15
16
16
10
15
15
12
11
10
9
K’
+1
10
8
5
5
3
0
5
5
4
4
5
5
3
5
0
1
5
2
4
2
6
6
4
3
0
1
0
1
2
0
0
3
3
0
4
5
5
K
-1
’
4
5
J’
15
16
17
17
15
15
15
15
16
16
8
14
14
15
16
16
15
14
14
14
14
13
13
13
15
8
8
15
18
7
7
15
9
16
9
9
10
J’’
14
15
16
16
14
14
14
14
15
15
7
13
13
14
15
15
14
13
13
13
13
12
12
12
14
7
7
14
17
6
6
14
8
8
9
15
8
 obs
7973.81400
7977.91480
8265.73370
8302.35480
8305.31120
8454.27510
8587.26500
8591.03200
8612.29900
8694.26730
8302.35480
8843.46960
8844.42330
8845.13850
8846.09220
8886.52800
8913.66000
9188.03220
9190.89320
9196.13840
9233.66550
9254.50330
9295.98810
9384.15530
9385.63350
9438.03790
9440.89890
9491.01450
9491.63440
9544.75400
9924.60250
10095.35790
10097.55130
10156.34540
10696.12500
10702.75310
11290.59790
4
4
3
1
4
4
4
3
4
4
3
4
1
0
5
1
4
2
6
6
3
3
0
0
1
1
2
1
0
3
3
1
4
5
5
K
-1
’’ K
+1
’’
4
5
9
7
3
4
11
17
2
3
10
5
5
5
12
4
9
14
16
16
9
8
11
12
15
15
4
10
11
14
15
15
9
14
14
11
10
9
8

Parameter
A /MHz
B /MHz
C /MHz
 μ
 μ
 μ c a b

/D

/D

/D
14 N(1)

/MHz

 aa bb cc
/MHz
/MHz
14 N(3)
 aa
/MHz
 bb
 cc
/MHz
/MHz
N c
ΔE /cm -1
EXPERIMENT
Rotamer
886.0063 (24) d
335.59351 (71)
270.10754 (36)
THEORY
NOT CONCLUSIVE IDENTIFICATION Anti/C2’endo t
Syn/C3’endo g+
901.2
935.8
790.0
799.7
925.5
340.6
276.6
308.4
266.6
352.6
261.4
330.6
262.9
300.4
264.0
Observed
Observed
Observed
2.1
3.7
1.6
2.8
3.1
1.5
0.9
0.7
0.8
4.5
0.9
1.3
2.3
2.3
0.2
-
-
-
66
-
-
-
-
1.50
1.43
-2.93
1.74
1.11
-2.85
-
0
1.82
0.73
-2.56
2.03
0.47
-2.50
-
383
1.48
1.71
-3.19
1.78
1.34
-3.12
-
664
1.46
1.81
-3.27
1.62
1.51
-3.13
-
701
1.82
-0.72
-1.11
1.98
-0.75
-1.23
-
751
1 Ab initio calculations MP2-6-311++G(d,p) below 1000 cm -1

• 3-10GHz
• Third harmonic of a 20 picosecond laser
• Microwave radiation pulse of 0.3
 s duration
• Frequency accuracy better than 5 kHz and an estimated resolution of 7 kHz
FT-MW
10
19
6075
 /MHz
6080

9
18
Broadband CP-FTMW rotational transition
Fabry-Pérot Resonator
Picosecond Laser
LA-MB-FTMW at GEM, Valladolid
I.
Peña et al. JACS 134 (2012) 2305 – 2312
6079.5
2,9 2,8
2,8 2,7
1,10 1,9
0,10 0,9
6080.0
 / MHz
1,11 1,10
O
NH
14 N
2,10 2,9
6080.5
HO
H
H
OH
O
H
OH
H
N
14 N
O
Narrowband LA-MB-FTMW
Sub-Doppler resolution

Parameter
A /MHz
B /MHz
C /MHz
D
 μ
J
/kHz
 μ b a

/D
 μ c

/D

/D
14 N(1)

/MHz

 aa bb cc
/MHz
/MHz
14 N(3)
 aa
/MHz
 bb
 cc
/MHz
/MHz
N c
ΔE /cm -1
EXPERIMENT
Rotamer
885.98961 (14) d
335.59720 (35)
270.11270 (20)
0.0115 (10)
Observed
Observed
Observed
THEORY
Anti/C2’endo g+ 1 Syn/C2’endo g+
Anti/C3’endo g+ Anti/C2’endo t
Syn/C3’endo g+
901.2
340.6
276.6
-
2.1
3.7
0.9
935.8
308.4
266.6
-
1.6
2.8
0.7
790.0
352.6
261.4
-
3.1
1.5
799.7
330.6
262.9
-
4.5
0.9
1.3
925.5
300.4
264.0
-
2.3
2.3
0.2
1.540 (42)
1.456 (72)
-2.996 (72)
1.719 (40)
1.261 (70)
-2.979 (70)
47
1.50
1.43
-2.93
1.74
1.11
-2.85
0
1.82
0.73
-2.56
2.03
0.47
-2.50
383
1.48
1.71
-3.19
1.78
1.34
-3.12
664
1.46
1.81
-3.27
1.62
1.51
-3.13
701
1.82
-0.72
-1.11
1.98
-0.75
-1.23
751
1 Ab initio calculations MP2-6-311++G(d,p) below 1000 cm -1

1. Distances in Å
2.97
Intramolecular hydrogen bonding network: 1
O3  H  O2  H  O2
Weak interactions:
C2  H  O5 
C6  H  O5 
Anti/C2’ -endo g+

 Uridine has been placed in the gas phase by laser ablation and the most stable Anti/C2 ’ -endo g+ has been characterized by broadband CP-FTMW and narrowband LA-MB-FTMW
 The observation of this conformer is in accordance with ab initio results, but in relative contrast with those obtained by X-ray 1 where C3 ’endo/anti conformation has been observed. It has been shown that the C3 ’ -endo configuration, which is in RNA, is favored when an explicit water molecule is introduced into the calculation. Nevertheless, previous calculations indicate that the change of the sugar pucker from
C3 ’ -endo to C2 ’ -endo in single-stranded RNA is energetically possible
 The preference for furanoses in RNA might be due to the observed intramolecular hydrogen bond network and the weak interactions between uracil and ribose, which over stabilizes this species and prevents the generation of pyranoses.
1. E. A. Green et al., Acta Cryst. (1975), B31, 102

Grupo de Espectroscopia Molecular (GEM)
Laboratorios de Espectroscopia y
Bioespectroscopia, Unidad Asociada CSIC,
UVa,Valladolid, Spain
Grants CTQ 2010- 19008,
AYA 2009-07304 and AYA 2012-32032
CSD 2009-00038 Molecular Astrophysics
Grants VA070A08 and CIP13/01