Proto-Urea-RNA (W?hler RNA) Containing Unusually Stable Urea Nucleosides

Article abstract of DOI:10.1002/anie.201911746

The RNA world hypothesis assumes that life on Earth began with nucleotides that formed information-carrying RNA oligomers able to self-replicate. Prebiotic reactions leading to the contemporary nucleosides are now known, but their execution often requires specific starting materials and lengthy reaction sequences. It was therefore proposed that the RNA world was likely proceeded by a proto-RNA world constructed from molecules that were likely present on the early Earth in greater abundance. Herein, we show that the prebiotic starting molecules bis-urea (biuret) and tris-urea (triuret) are able to directly react with ribose. The urea-ribosides are remarkably stable because they are held together by a network of intramolecular, bifurcated hydrogen bonds. This even allowed the synthesis of phosphoramidite building blocks and incorporation of the units into RNA. Investigations of the nucleotides’ base-pairing potential showed that triuret:G RNA base pairs closely resemble U:G wobble base pairs. Based on the probable abundance of urea on the early Earth, we postulate that urea-containing RNA bases are good candidates for a proto-RNA world.

Full text of DOI:10.1002/anie.201911746

A Journal of the Gesellschaft Deutscher Chemiker
Angewandte
Chemie
International Edition
www.angewandte.org
Accepted Article
Title: proto-Urea-RNA (Wöhler RNA) containing unusually stable urea
nucleosides
Authors: Hidenori Okamura, Antony Crisp, Sarah Hübner, Sidney
Becker, Petra Rovo, and Thomas Carell
This manuscript has been accepted after peer review and appears as an
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To be cited as: Angew. Chem. Int. Ed. 10.1002/anie.201911746
Angew. Chem. 10.1002/ange.201911746
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http://dx.doi.org/10.1002/ange.201911746

10.1002/anie.201911746
Angewandte Chemie International Edition
RESEARCH ARTICLE
proto-Urea-RNA (Wöhler RNA) containing unusually stable urea
nucleosides
Hidenori Okamura,# Antony Crisp,# Sarah Hübner, Sidney Becker, Petra Rovó* and Thomas Carell*
Abstract: The RNA world hypothesis assumes that life on Earth
began with nucleotides that formed information carrying RNA
oligomers able to self-replicate. Prebiotic reactions leading to the
contemporary nucleosides are now known, but their execution often
requires specific starting materials and lengthy reaction sequences. It
was therefore proposed that the RNA-world was likely proceeded by
a proto-RNA world constructed from molecules that were likely
present on the early Earth in greater abundance. Here we show that
the prebiotic starting molecules bis-urea (biuret) and tris-urea (triuret)
are able to directly react with ribose. The urea-ribosides are
remarkably stable, because they are held together by a network of
intramolecular, bifurcated H-bonds. This even allowed the synthesis
of phosphoramidite building blocks and incorporation of the units into
RNA. Investigation of the nucleotides’ base pairing potential showed
replicate and which featured properties leading to their survival
under early Earth conditions.[14-17] It is assumed that based on the
processes of chemical evolution, more and more complex RNA
and RNA-peptide structures were created that finally led to the
emergence of life.[18-19] RNA and the constituting nucleosides that
are needed to establish faithful replication of “genetic” information
are, however, rather complex chemical structures. The problem
of finding prebiotically-plausible pathways to the canonical
nucleosides (Fig. 1 a) (known as the nucleoside problem)[20] led
to the idea that RNA was potentially proceeded by a proto-RNA
that could more easily arise from prebiotically privileged starting
materials.[21] As a result, emerging discussions about the origin of
life have often emphasised the significance of informational
polymers that are simpler than RNA. A revolutionary study from
that triuret:G RNA base pairs closely resemble U:G wobble base pairs. Eschenmoser’s group, for example, demonstrated that α-
Based on the probable abundance of urea on the early Earth, we
postulate that urea-containing RNA bases are good candidates for a
proto-RNA world.
threofuranosyl nucleic acid (TNA) is capable for forming
antiparallel duplexes and can even pair with cDNA or RNA.[22]
TNA was later simplified to an acyclic polymer known as glycol
nucleic acid (GNA)[23-25] and various other XNA backbones have
since been investigated.[26] We know that formaldehyde and
Ca(OH)2 can give sugars via the formose reaction,[27-29] as was
first described by Butlerov[30]. Though it remains unclear how
exactly such a process could lead to the substantial accumulation
of ribose on the early Earth, recent developments have led to
significant improvements in prebiotic the synthesis of ribose from
simpler aldoses.[31-33] We also know that urea was one of the most
likely nitrogen-containing molecules present on the early Earth
and that it can activate mineralic phosphate to achieve
phosphorylations.[4-5] Because urea itself has even been shown to
react directly with ribose under mild acidic conditions,[34] we asked
whether the construction of informational base pairing systems
based on the pyrolysis products of urea, biuret and triuret (Fig.
1b),[35] would be possible.
Introduction
Urea, the bisamide of carbonic acid, is widely distributed in the
biosphere and plays a fundamentally important role in the
biosynthesis of proteins and the entire N-cycle of organisms in
general.[1-2] It is also believed to have formed early on the prebiotic
Earth and before the process of chemical evolution that gave the
centrally important molecules of life.[3] Urea is a key starting
molecule for many prebiotic chemical reactions,[4-11] and was the
first organic compound synthesized from inorganic matter
(ammonium cyanate) by the chemist Friederich Wöhler in 1828.[12]
Wöhler’s synthesis gave birth to the field of organic chemistry and
was among others, essential to defeating the mainstream
ideology of vitalism, which stated that organic matter contained a
special vital force.[13] Current theories about the origin of life build
upon the RNA world hypothesis, which predicts the early
formation of information-encoding RNA that was able to self-
(a) canonical RNA nucelosides
O
NH2
N
NH2
O
N
N
NH
O
O
O
O
O
N
N
N
N
N
NH
NH2
O
HO
HO
HO
HO
N
N
OH
HO
OH
OH
OH
HO
HO
HO
A
U
C
G
(c) Wöhler bases
O
(b) polyureas
HN
H
O
H
N
[a]
Dr. Hidenori Okamura, B. Sc. Antony Crisp, B. Sc. Sarah Hübner,
Dr. Sidney Becker, Dr. Petra Rovó and Prof. Dr. T. Carell
Center for Integrated Protein Science (CiPSM) at the Department of
Chemistry, LMU München
Butenandtstr. 5-13, 81377 München
Fax: (+) 49 2180 77756
NH2
O
O
O
NH
NH
O
H-bond
donors &
acceptors
biuret
triuret
H
NH
H2N
O
N
H
NH2
O
O
N
O
O
HO
O
HO
OH
HO
H2N
N
H
N
H
NH2
OH
HO
β-1
β-2
E-mail: petra.rovo@lmu.de and thomas.carell@lmu.de
Homepage: www.carellgroup.de
Figure 1. (a) The chemical structures of the canonical RNA nucleosides. (b)
The chemical structures of biuret and triuret. (c) Depiction of the urea based
nucleosides with potentially stabilizing H-bonds.
# these authors contributed equally.
Supporting information for this article is given via a link at the end of
the document.
This article is protected by copyright. All rights reserved.

10.1002/anie.201911746
Angewandte Chemie International Edition
RESEARCH ARTICLE
The fundamental chemical problem associated with this notion is
that such nucleosides (Fig. 1c) should be highly prone to
hydrolysis in an aqueous environment. Investigating the
structures of potential urea bases, however, led us to discover
that those containing biuret (b-1) and triuret (b-2) are highly
stabile even in water, probably due to the intramolecular H-bonds
that they provide (Fig. 1c). The question of the possible prebiotic
existence of urea (Wöhler) RNA is therefore directly associated
with the question of to which extent these non-covalent
interactions protect from hydrolysis.
The mixture was subsequently analyzed by reverse phase HPLC-
MS. Gratifyingly, we noted significant formation of the
corresponding nucleosides with both biuret and triuret. These
wet-dry conditions mimic the intermittently concentrating
environments that might be found on drying beaches or lagoons
(or even today in Death Valley),[42] as was recently discussed in
relation to the prebiotic synthesis of canonical and non-canonical
purine nucleosides.[39] For the biuret reaction, we detected
formation of four major nucleoside products. Of those, two are the
a- and b-anomers of the ribofuranosides ( -1 and -1). These
a b
structures were confirmed by independent synthesis of the a- and
b-anomers (vide infra) followed by co-injection studies. The
absolute stereochemical configurations of the a- and b- anomers
were confirmed by NOESY-NMR spectroscopy (SI). The other
two compounds detected in the HPLC-MS experiment are likely
the pyranosidic species (not further investigated). Similar but not
identical data were obtained for triuret. Here too, we detected four
reaction products of which two are the a- and b-anomers of the
Results and Discussion
In order to investigate the formation of urea nucleosides under
plausible prebiotic conditions, we mixed aqueous solutions of
ribose with either biuret or triuret in the presence of boric acid and
heated the mixture at 95 °C for 18h, allowing the mixture to slowly
dry down. Boric acid, which due to its high electronic deficiency
forms complexes with vicinal 1,2-diols, was included for its known
stabilizing[36-38] and directing[39-41] effects on ribose. The resulting
solid was then taken up in dilute (100 mM) sodium carbonate
buffer (pH = 9.5) and heated again at 95ºC for 1h (Fig. 2).
ribofuranosides ( -2 and -2), with the remaining compounds
a b
likely being again the pyranosides. We focused our initial studies
on the ribofuranosides and noted to our surprise, that both the
biuret and the triuret species are quite stable. Both compounds
can be kept for prolonged periods in an aqueous solution at
neutral pH without signs of anomerisation. This unusual stability
of the biuret and triuret nucleosides, prompted us to study if one
could generate phosphoramidites and insert them into RNA. This
would require that the Wöhler nucleosides survive even the
nucleophilic reaction and deprotection conditions needed for solid
phase RNA synthesis.
NH2
NH2
(a)
i. Ribose, Boric acid,
95 ºC, dry-state
ii. NaHCO3(aq)
O
H
N
O
H
N
NH
O
O
O
NH
O
O
O
+
H2N
N
NH2
HO
HO
H
OH
HO
OH
HO
-1
-1
100
75
50
25
0
100
75
50
25
0
MS
UV
-1
-1
-1
-1
The synthesis of the phosphoramidite building blocks is shown in
Scheme 1. We began by 3’,5’-silyl-protecting 1-azidoribose 3 to
obtain 4, followed by TOM-protection of the 2’-OH group to give
5. Desilylation of the 3’,5’-positions (affording compound 6) and
subsequent DMTr protection of the 5’-OH group furnished
compound 7. We then protected the 3’-OH group with an acetyl
group to give 8 and reduced the azide by catalytic hydrogenation
followed by reaction of the amine with trimethylsilylisocyanate.
This provides the urea riboside 9. A second reaction with
trichloroacetylisocyanate in pyridine followed by cleavage of the
trichloroacetate group with basic alumina in methanol gave the
4
5
6
7
4
5
6
7
Time (min)
Time (min)
O
O
(b)
HN
H
HN
H
O
H
iii. Ribose, Boric acid,
95 ºC, dry-state
iv. NaHCO3(aq)
NH
NH
NH
NH
O
O
O
O
O
+
H
H2N
N
N
NH2
biuret riboside 10 as a mixture of the a- and b-isomers (
-10) which we separated via column chromatography (SiO2,
CH2Cl2:CH3OH = 100:0 → 98:2). The unwanted a-isomer ( -10)
was able to be isomerized upon heating in the presence of DBU,
which gave a mixture of -10 and -10. Iterative anomerisation
allowed us to increase the total isolated yield of the b-anomer (
b
-10 and
O
N
O
N
H
H
O
HO
a
HO
OH
HO
a
OH
HO
-2
-2
100
75
50
25
0
100
MS
UV
-2
b
a
-2
b
-
-2
-2
75
50
25
0
10) for subsequent reactions. To obtain the triuret nucleoside 11,
we repeated the trichloroacetylisocyanate reaction followed by
basic alumina treatment using the pure b-anomer of the biuret
nucleoside
( -10). Interestingly, we observed very little
b
anomerisation during these reactions (as monitored via TLC). The
b-triuret nucleoside 11 was therefore obtained in anomerically
b
pure form and in 58% yield. Both the b-biuret ( -10) and the b-
9
10
11
12
13
9
10
11
12
13
Time (min)
Time (min)
triuret (11) nucleosides were next converted into the
corresponding phosphoramidites. To this end we cleaved the 3’-
acetyl group with ammonia (even this does not lead to hydrolysis
Figure 2. Reaction of (a) biuret and (b) triuret with ribose and analysis of the
reaction mixture by HPLC-MS, showing successful formation of urea-based
nucleosides.
This article is protected by copyright. All rights reserved.

10.1002/anie.201911746
Angewandte Chemie International Edition
RESEARCH ARTICLE
of the glycosidic bond), and then treated the ensuing compounds
12 and 13 with bis(2-cyanoethyl)-N,N-diisopropylphosphor-
amidite in the presence of diisopropylamine and tetrazole. This
final step provided the phosphoramidite building-blocks 14 and 15
in fair yields of 70% and 54% respectively.
efficiently produced and can be cleanly purified. Similar data were
obtained for the biuret containing strands (Fig. S7). It is a
remarkable result that the urea bases survive the RNA synthesis
conditions to give RNA strands in excellent purity.
O
NH2
O
O
N3
O
O
N3
OH
N3
OH
NH2
O
a
b
O
O
NH
HO
O
t-Bu
i. RNA synthesis (ultramild)
ii. NH3/MeOH, r.t.
iii. 3HF.TEA, TEA,
DMSO, 65 ºC
Si
t-Bu
NH
t-Bu
OTOM
O
O
Si
t-Bu
NH
O
HO
O
DMTrO
NH
O
O
O
5
4
NH
3
O
O
NH
OH
OTOM
CN
O
P
O
P
N
O
-
O
O
O
N3
e
N3
d
N3
c
-
O
HO
DMTrO
DMTrO
OTOM
raw-HPLC
HO
1800
MALDI-
TOF MS
OTOM
AcO
OTOM
HO
3.0
2.0
1.0
6
8
7
1400
1000
600
O
NH2
O
[M–H]–: 3113.08
calc: 3113.41
O
NH
NH2
O
NH
OTOM
200
0
O
NH
0
f, g
h, i
DMTrO
DMTrO
0
10
20
30
40
2000 4000
m/z
0
6000
AcO
Time (min)
OTOM
AcO
α-10
β-10
9
Figure 3. Preparatory HPLC and MALDI-TOF mass data for an example Tri
containing RNA strand with the sequence 5’-CUUACTriCUGA-3’.
j
O
NH2
This stability allowed us next to investigate the pairing properties
of the biuret and triuret bases using thermal melting curve studies
(10 mM Na-phosphate, 150 mM NaCl, pH 7.0). If a candidate
proto-RNA were to have existed before the emergence of modern
RNA, then it stands to reason its bases needed to pair with the
canonical nucleosides to allow a smooth evolutionary transition
from proto-RNA to RNA. For the measurements we prepared
RNA strands with a C:G or a U:A base pair in a central position.
We then exchanged the pyrimidine base C or U with the biuret
(Bi) or triuret (Tri) base. The data are compiled in Table 4b. The
unmodified RNA duplexes feature, as expected, rather high
melting temperatures of 55 °C (C:G) and 49 °C (U:A). For all
Bi:A/G/U/C base pairs we measured much lower melting
temperatures, which were in addition, quite similar irrespective of
the counter base (between 28 °C and 31 °C). This shows that the
biuret base does not prefer a particular counter base and that
base pairing is in general weak, if it occurs at all. For the larger
triuret base we noted significantly higher melting temperatures
between 35 °C and 45 °C. In addition we observed a clear base
pairing preference with G. The melting temperature for the Tri:G
base pair is around 5 °C higher than the others, which clearly
points to a significant pairing selectivity. Analysis of the base
pairing potential of the triuret base shows that it is in principle
possible to form a wobble type base pair with G. We therefore
tested an RNA duplex with a central U:G wobble base pair and
observed indeed the same melting temperature (45 °C). Our
hypothesis was further validated by the observation that triuret
also forms a stable base-pair with the structurally related base
inosine (I), giving an almost identical melting temperature of 44 °C.
To exclude that this is pure chance, we next prepared an RNA
strand with either two or even three consecutive triuret bases and
paired this strand with a counter strand containing either two or
three central G bases. Comparative melting point studies show
exactly the same behavior between the U:G wobble and the Tri:G
base pairs.
O
NH
O
O
NH2
NH2
O
NH
O
O
NH
m
o
NH
DMTrO
O
O
NH
NH
OTOM
CN
O
DMTrO
DMTrO
P
N
O
OTOM
HO
OTOM
β-10
AcO
12
14
O
NH2
k, l
O
O
O
O
NH2
NH2
NH
O
O
NH
NH
NH
O
O
n
p
O
NH
NH
NH
DMTrO
N
O
O
NH
OTOM
NH
DMTrO
DMTrO
OTOM
CN
O
P
HO
OTOM
11
AcO
O
13
15
Scheme 1. Phosphoramidite building block synthesis of the biuret and triuret
nucleosides. Conditions: a. t-Bu2Si(OTf)2, DMF, 0 °C, 1 h. b. i-PrSiO(CH2)Cl,
NaH, THF, 0 °C, overnight, 55% over two steps. c. HF-pyridine, pyridine, CH2Cl2,
r.t., 1 h, 61%. d. DMTrCl, pyridine, r.t., overnight, 77%. e. Ac2O, DMAP, pyridine,
r.t., 2 h, 91%. f. 10% Pd/C, H2, THF, r.t., 2 h, then g. TMS-isocyanate, THF, r.t.,
overnight, 76% (mixture of diastereomers). h. trichloroacetylisocyanate,
pyridine, THF, r.t., 1h, then i. Al2O3, MeOH, r.t., 1 h, 90% (dr a:b = 1.6:1). j. DBU,
THF, 50 °C, overnight, 28% (96% brsm). k. trichloroacetylisocyanate, pyridine,
THF, r.t., 1 h, then l. Al2O3, MeOH, r.t., 1 h, 58%. m. NH3, MeOH, r.t., 4 h, 74%.
n.
NH3,
MeOH,
r.t.,
4
h,
76%.
o.
bis(2-cyanoethyl)-N,N-
diisopropylphosphoramidite, diisopropylamine-tetrazole, CH3CN, r.t., overnight,
70%. p. bis(2-cyanoethyl)-N,N-diisopropylphosphoramidite, diisopropylamine-
tetrazole, CH3CN, r.t., overnight, 54%.
For the solid phase oligonucleotide synthesis, we used a standard
ultramild RNA synthesis protocol. The urea bases were coupled
once for 20 min. After full assembly of the RNA strands using Pac-
chemistry conditions, we deprotected the RNA strands and
cleaved them from the solid support with NH3 at r.t. in methanol.
The silyl protecting groups were finally removed using HF-TEA in
DMSO at 65 °C. Fig. 3 shows the raw HPLC chromatogram of the
triuret containing RNA strand as an example, as well as the
MALDI-TOF mass spectrum obtained from purification of the
major species. Clearly evident is that the RNA strands are
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10.1002/anie.201911746
Angewandte Chemie International Edition
RESEARCH ARTICLE
Wöhler RNA
base-pair
U–G wobble
base-pair
(a)
peak with H5 (purple line in Fig. 5c). These, as well as a handful
other NOE cross peaks between the triuret base protons and the
surrounding protons support the structural model depicted in Fig.
4a (all other NOE distance restraints are depicted with black lines
in Fig. 5b), with the triuret moiety clearly forming bifurcated
hydrogen bonds.
O
O
HN
H
O
H
NH
N
N
O
H
O
N
N
N
H
O
N
N
N
RNA
H
N
RNA
N
N
RNA
O
H
RNA
H2N
N
H2N
(b)
!"#$%&'&(!)& !"#$%&*&(")&
!
+&(,-)&
//&&
(c)
-&
0&
0&
.&
1&
.&
50.0
40.0
30.0
20.0
10.0
0.0
23&&
#$%%
4#&
4#&
1&
.&
0&
-&
1&
.&
0&
-&
:&
*5&&
*3&&
6'&&
67&&
27&&
#$%
4#&
4#&
89#&
89#&
89#&
89#&
89#&
ii
i
U-A
U-G
Tri-G
63&&
6/&&
22&
5'-CUUAXCXUGA-3'
3'-GAAUYGYACU-5'
i
5'-CUUAXXXUGA-3'
3'-GAAUYYYACU-5'
Oligo 1: 5'-CUUACXCUGA-3'
Oligo 2 : 3'-GAAUGYGACU-5'
ii
Figure 4. (a) Chemical structures and base-pairing properties of triuret and
similarity between the Tri:G base pair and a U:G wobble base-pair. (b) Summary
of Tm analyses for oligonucleotides containing biuret and triuret. (c) Summary
of Tm analyses for oligonucleotides containing more than one modified-base or
U:G wobble-base pair. Solutions were buffered with 10 mM Na-phosphate (pH
7.0) and 150 mM NaCl.
To gain deeper insight into the origin of the stability of the Tri:G
base pairing we prepared 8 mer palindromic RNA strands which
are designed to form canonical C:G, wobble U:G or Tri:G pairs at
the central position (GGUXGACC, where X = C, U, or Tri) and
1
analyzed their 2D H-1H NOESY spectra.[43] The high chemical
shift similarity in the fingerprint region of the spectra (Fig. S17)
confirms the same overall structure for the three oligonucleotides
namely formation of an A-form double-stranded RNA (Fig. 5c).
The imino region of the NOESY spectra gives direct indication
about the H-bond interactions between base pairs (Fig. 5a, Fig.
S16). The Tri base has five amide protons (H1, H3, H5, H71, and
H72) and three carbonyl oxygens that could potentially be
involved in base pairing. Out of the five protons, H3 and H5 are
partially or fully solvent exposed and thus they show strong
exchange cross peaks at the water resonance (Fig. S19). The
number and intensity of cross-peaks between the H1 proton of Tri
and the sugar/base protons of the previous uridine base (U3)
indicate that the H1 proton is located in a similar arrangement to
the H5 proton of a U or a C or as the H8 proton of an A and a G
base in canonical dsRNA structures; e.g. we observed a strong
NOE cross peak between Tri4H1 and U3H2’ (yellow line in Fig.
5b). Similarly, the terminal NH2 group of Tri shows strong cross
peaks with the H2’ and H5 protons of U3 (green lines in Fig. 5c).
Thus, the NOESY spectra suggest both H1 and the NH2 group
point towards the phosphate backbone. Direct evidence for Tri:G
base pairing was obtained from a NOESY spectrum recorded with
relatively short (40 ms) mixing time (Fig. 5a). This experiment
provides a clear cross peak between the H1 imino proton of G5
and the H3 amide proton of Tri of the opposite strand (red line in
Fig. 5b). In addition, the TriH3 proton shows an intrabase cross
Figure 5. NMR analysis of the Tri:G base pairing. (a) Excerpt from the 1H-1H
NOESY spectrum (tmix = 40 ms) of the dsRNA (GGUXGACC, where X = Tri)
showing the inter-strand cross peaks between the H1 imino proton of G5 and
the H3 amide proton of Tri4. Inset shows a model for the Tri:G base paring. (b)
NOE connections of the triuret base amide protons. Essential structure defining
NOE connections are highlighted for Tri4H1 – U3H2’ (yellow), Tri4H71/H72 –
U3H2’ (green), G5H1 – Tri4H3 (red), Tri4H3 – Tri4H5 (purple). Other observed
NOE connections are displayed with black lines. (c) Structural model of the
GGUXGACC oligonucleotide showing the non-canonical base pairing between
G5 (blue) and Tri4 (green) bases.
To obtain a three-dimensional model for the Tri:G base pair, we
performed molecular modeling using the online software
ROSIE[44-45] followed by NOE-based structure calculation using
the software CNSsolve.[46-47] During the structure calculation, only
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10.1002/anie.201911746
Angewandte Chemie International Edition
RESEARCH ARTICLE
the conformation of the triuret base was altered, while the
phosphate backbone and all other bases were kept at their fixed
position. Fig. 5c displays the obtained low energy structure model
for the double-stranded 8mer RNA. Clearly evident is that the two
Tri:G base pairs are well accommodated in the structure and that
the extended network of H-bonds within the Tri structure and
between Tri and the opposite G establish the measured stability.
obtained upon pyrolysis of urea, one of the most likely building
blocks available on the early Earth. Given that various tri- tetra-
and pentose sugars are prebiotically accessible from
glycoaldehyde,[31-32] which is itself accessible from either
formaldehyde[30] or HCN by ultraviolet irradiation,[49] our discovery
creates the prebiotically attractive possibility of generating
information encoding oligomers whose key building blocks are
derived of simple 1-carbon units. The chemistry described here
now needs to be explored with sugars simpler than ribose.
All these results confirm that triuret, which is itself a condensation
product of urea, is able to form a folded pseudobase that pairs
with guanine. Finally, in order to further demonstrate the prebiotic
plausibility of Wöhler RNA, we synthesized a homo-RNA-
oligomer containing exclusively triuret bases (Fig 6a). The only
additional structural modification was the inclusion of a dye at the
5’-end (Cy3), which was necessary to allow UV detection at 548
nm and therefore purification via HPLC. Remarkably, despite
having five, in principle hydrolysable triuret bases in a row, the
homo-strand was bench stable both at room temperature, as well
as when subjected to the harsh conditions necessary for RNA
deprotection and cleavage from the solid support. Fig. 6 shows
the raw-HPLC chromatogram obtained from the material directly
after synthesis and the correct MALDI-TOF mass spectrum
verifying the integrity of the material.
50]
25]
Discussed examples are threose-[22,
backbones.
and glycol-[23,
based
Acknowledgements
We thank the Deutsche Forschungsgemeinschaft for financial
support via SFB1309, SPP1784 and CA275/11-1. This project
has received funding from the European Research Council (ERC)
under the European Union's Horizon 2020 research and
innovation programme (grant agreement n° EPiR 741912) and
through a H2020 Marie Skłodowska-Curie Action (LightDyNAmics,
765866). H.O. acknowledges support from Marie Sklodowska
Curie Individual Fellowship (752420) from the European
Commission. P.R. acknowledges support from the Center for
NanoScience and from the Fonds der Chemischen Industrie.
(a)
(b)
100
75
raw-HPLC
R = Pr-Cy3+-Pr-OH
H
N
O
OR
H
O
O
P
O-
NH
NH
H
N
O
Keywords: prebiotic chemistry • origin of life • urea • proto RNA
50
O
• base pairing •
H
N
O
O
O
P
OH
O-
25
H
O
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NH
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(c)
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[M–2H]–: 2142.33
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RESEARCH ARTICLE
Entry for the Table of Contents
RESEARCH ARTICLE
Hidenori Okamura, # Antony Crisp, #
Sarah Hübner, Sidney Becker, Petra
Rovó,* and Thomas Carell.*
Page No. – Page No.
proto-Urea-RNA (Wöhler RNA)
containing unusually stable urea
nucleosides
Formation of bis-urea (biuret) and tris-urea (triuret) nucleosides under prebiotically plausible conditions provides the foundation for a
novel proto-RNA concept.
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