Solid Supports for the Synthesis of 3′-Aminooxy Deoxy- or Ribo-oligonucleotides and Their 3′-Conjugation by Oxime Ligation

Article abstract of DOI:10.1021/acs.joc.9b00848

Mono- and triethylene glycol aminooxy derivatives were reacted with levulinic acid, protected with dimethoxytrityl, and immobilized on solid support. The resulting solid supports were used for elongation of oligonucleotides. Then, a mild ammonia treatment was applied to remove the oligonucleotide protecting groups, followed by a treatment with 50 mM methoxyamine at pH 4.2, releasing the 3′-aminooxy oligonucleotides by an oxime exchange reaction. The resulting 3′-aminooxy deoxy- or ribo-oligonucleotides were conjugated to various ketones and aldehydes with high efficiency by oxime ligation.

Full text of DOI:10.1021/acs.joc.9b00848

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Note
Solid supports for the synthesis of 3'-aminooxy deoxy- or ribo-
oligonucleotides and their 3'-conjugation by oxime ligation
Mathieu Noel, Celine Clement-Blanc, Albert Meyer, Jean-Jacques Vasseur, and Francois Morvan
J. Org. Chem., Just Accepted Manuscript • DOI: 10.1021/acs.joc.9b00848 • Publication Date (Web): 15 Oct 2019
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Solid supports for the synthesis of 3'-aminooxy deoxy- or ribo-
oligonucleotides and their 3'-conjugation by oxime ligation
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Mathieu Noël, Céline Clément-Blanc, Albert Meyer, Jean-Jacques Vasseur and François
Morvan*
Institut des Biomolécules Max Mousseron, Université de Montpellier, CNRS, ENSCM,
Montpellier, France
Graphical Abstract
Oligonucleotide
N
N
O
synthesis
DMTr
O
NH
NH
n
n
O
O
O
O
n = 1 or 3
NH4OH
+
MeONH2 NH4 AcO- pH 4.2
H
R
O
O
n
N
O
n
NH2
O
O
+
NH4 AcO-
R
pH 4.2
ABSTRACT
Mono- and tri-ethyleneglycol aminooxy derivatives were reacted with levulinic acid, protected
with dimethoxytrityl and immobilized on solid support. The resulting solid supports were used
for elongation of oligonucleotides. Then, a mild ammonia treatment was applied to remove the
oligonucleotide protecting groups, followed by a treatment with 50 mM methoxyamine at pH
4.2, releasing the 3'-aminooxy oligonucleotides by an oxime exchange reaction. The resulting
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3'-aminooxy deoxy- or ribo-oligonucleotides were conjugated to various ketones and aldehydes
with high efficiency by oxime ligation.
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Keywords: Oligonucleotides, Oxime, solid support, click, conjugates
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Conjugation of oligonucleotides to diverse molecules has attracted substantial research interest
as it offers an easy way to modulate their biological properties. Most of the conjugations are
made either at the 5' or the 3' extremity of the oligonucleotides. The 3'-modification has a great
interest as it also increases the nuclease resistance.1
Different chemistries have been applied to conjugate oligonucleotides with different
biomolecules or dyes including Diels-Alder ligation,2,3 Staudinger ligation4 copper catalyzed
10-13
alkyne–azide cycloaddition,5-9 copper free alkyne–azide cycloaddition,
thiol Michael
addition,14-18 thioether ligation,19 inverse-electron-demand Diels–Alder reaction20,21 or Oxime
ligation.22-30
With regards to oxime ligation, several phosphoramidite derivatives have been reported for the
introduction of the aminooxy function at the 5'-end of oligonucleotides for their subsequent
conjugation or cyclization.23-25,31-35 In contrast, the 3'-conjugation of oligonucleotides by oxime
ligation has been poorly explored. The usual method reported in the literature to do so is to
introduce a glycerol group at the 3'-end which is further oxidize to an aldehyde to eventually
reacts with a aminooxy derivative. Thus, the modulation of the spacer between the
oligonucleotide and the modification is not possible. Only one recent publication, from our
group, reported the synthesis of a solid support allowing for the introduction of the aminooxy
function at the 3'-end of oligonucleotides (Figure 1).35 The resulting 3'-modified
oligonucleotides were then conjugated with different aldehyde derivatives or circularized by
means of bis-oxime ligation after the introduction of another aminooxy function at the 5'-end
and reaction with a dialdehyde.35
In order to introduce some versatility by easily modulating the length of the spacer between an
oligonucleotide and its 3'-aminooxy function for subsequent oxime conjugation, we designed a
new type of solid supports built from ethylene glycol derivatives (Figure 1). Herein, we report
the synthesis of two solid supports allowing the 3'-conjugation of deoxy- and ribo-
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oligonucleotides with different ketones and aldehydes. This novel approach allows for
enhanced versatility in generating a variety of tailored oligonucleotide constructs, where the
different solid supports can be utilized to efficiently modulate the target designs of 3'-
oligonucleotide conjugates.
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DMTr
O
O
O
O
O
Previous work
Current work
N
O
N
H
O
O
N
DMTr
O
NH
n
O
O
7a
7b
n = 1
n = 3
Figure 1. Structure of previous phthalimide solid support35 and new solid supports 7a-b.
To demonstrate the usefulness of this approach, we selected mono- and tri-ethyleneglycol as
the starting compounds. However, it is worth noting that this synthetic strategy can be readily
applied to ethyleneglycols of any length or to any diols. The synthesis of the solid supports 7a-b
leading to 3'-aminooxy oligonucleotides with a mono- or a tri-ethylene glycol spacer proceeded
in few steps (Scheme 1). Firstly, 2-bromo-ethanol, or a previously synthesized monotosyl-
triethyleneglycol (1)36 were substituted with hydroxyl-phthalimide in the presence of K2CO3
affording the phthalimide ethyleneglycol derivatives 2a-b.23,35 Secondly, the aminooxy
function was deprotected with hydrazine, and after evaporation, levulinic acid was added to
form the oxime derivatives 5a-b. After work-up, the crude was directly treated with
dimethoxytrityl chloride to protect the hydroxyl function providing derivatives 6a-b after
chromatography with an overall yield of 50% for the last three steps. Finally, 6a-b were
immobilized by amide formation on long-chain alkylamino controlled-pore glass (LCAA-CPG
1000 Å), using EDC as the coupling reagent, affording the two solid supports 7a-b. The loading
was measured by trityl assay (~40 µmol/g).
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H
O
OH
3
5
6
Tos-Cl
O
7
8
9
Tos
O
N
OH
OH
3
O
NH2NH2
Pyridine
1
O
3a
3b
n = 1
n = 3
N
O
H2N
O
OH
n
OH
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n
K2CO3
DMF
Br
O
OH
2a
2b
n = 1
n = 3
O
AcONH4
OH
MeOH / DMF
O
4
NH2
EDC
Pyridine
Et3N
DMTr-Cl
Pyridine
N
N
O
N
O
DMTr
O
O
OH
n
O
DMTr
NH
n
n
O
HO
O
OH
O
7a
7b
6a
6b
5a
5b
n = 1
n = 3
n = 1
n = 3
n = 1
n = 3
Scheme 1. Synthesis of solid supports for the preparation of 3'-aminooxy oligonucleotides.
The elongation to full-length oligonucleotides (DNA or RNA) was performed on a DNA/RNA
synthesizer, at the 1 mol scale, by means of solid phase oligonucleotide synthesis (SPOS)
using standard 3'-phosphoramidite chemistry with fast-labile nucleobase protecting groups and
2'-O-pivaloyloxymethyl (PivOM) groups for the 2'-OH protection of the ribonucleotides,37
(Scheme 2). After elongation, the oligodeoxynucleotides 8a-b were first deprotected with
concentrated aqueous ammonia for 1 h at room temperature to remove the cyanoethyl groups
on the phosphates and the acyl groups on the nucleobases. For oligoribonucleotides a first-step
treatment with 1 M DBU was applied to remove the cyanoethyl groups and then the second-
step ammonia treatment for 3 h at 37 °C was performed to allow removal of the PivOM groups
on the 2' position of ribose.37 The ammonia solution was collected, and after evaporation, the
residue was analyzed confirming no undesired cleavage of the oligonucleotides from the solid
supports. Finally, the 3'-aminooxy-oligonucleotides 10a-b were released from the solid
supports by treatment with a 0.4 M ammonium acetate buffer solution of 50 mM methoxyamine
(pH 4.2) for 3 h at rt. The 3'-aminooxy-oligonucleotides were purified by size exclusion
chromatography on G25-sephadex (Nap-10) to remove the methoxyamine and the salts. The 3'-
aminooxy-oligonucleotides were obtained in the range of 300 to 500 nmoles that is a similar
amount in comparison with oligonucleotides synthesized on a standard succinyl solid support.
HPLC and MALDI-TOF MS analyses showed the efficiency of the syntheses and confirmed
the nature of the 3'-modified oligonucleotides (See SI). It has been reported that methoxyamine
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reacts with certain cytosines of tRNA. 38 Such degradation occurred after a 20 h treatment at 37
°C with a 3 M methoxyamine concentration and 10 mM MgCl2 pH 5.5. In our case, no similar
side reaction was observed, this could be explained by the short treatment (3 h) at room
temperature with a low concentration of methoxyamine (50 mM) applied for the release of the
aminooxy-oligonucleotides from the solid support.
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B*
HO
O
O
P
R
B*
O
O
O
SPOS
O
Cne
N
N
n
DMTr
O
n
O
R
O
NH
O
O
NH
O
P
m
O
Cne
O
O
7a
7b
8a
8b
n = 1
n = 3
n = 1
n = 3
R = H or O-CH2OPiv
1M DBU in CH3CN for RNA
aq NH4OH
H2O
B
HO
B
R
HO
O
O
O
CH3ONH2
O
P
O-
R
O
O
B
O
B
pH 4.2
NH4+ AcO-
O
P
O-
O
P
O
N
O
R
O
NH2
O
R
O
n
O
m
NH
O
m
n
-O
P
-O
R = H or OH
O
O
O
R = H or OH
10a
10b
9a
9b
n = 1
n = 3
n = 1
n = 3
Scheme 2: Synthesis of 3'-aminooxy-oligonucleotides. B* represents the protected nucleobase
APac, G-tbu-Pac, C-Ac. Cne: cyanoethyl, Piv: pivaloyl. SPOS: Solid phase oligonucleotide
synthesis.
To confirm the reactivity of the 3'-aminooxy-oligonucleotides, they were then engaged in
derivatization reactions using different ketones and aldehydes, like acetone, cyclohexanone,
fluorenone, pyrene carboxaldehyde, biotin or deoxycholic acetamide aldehydes (Scheme 4 and
table 1). The aldehyde derivatives of biotin and deoxycholic acid 14 and 18 were synthesized
in three steps, as follows: first, activation of the carboxylic function of biotin and deoxycholic
acid as a N-hydroxysuccinimide derivative; second, conjugation with 3-amino-2-hydroxy-
propan-1-ol; and, third, oxidation of the diol by treatment with sodium periodate to generate
the aldehyde function (Scheme 3).39
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5
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O
O
H2N
OH
NH
OH
NH
H
HN
H
HN
H
H
O
O
OH
7
8
9
S
HN
S
R
OH
CH3CN
Et3N
O
13
O
11
R = OH
12
MeOH
H2O
HO
N
10
11
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13
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Na IO4
R =
O
N
O
O
O
NH
H
DCC
DMF
HN
O
H
H
S
HN
O
14
OH
OH
H2N
OH
H
H
H
H
HO
OH
HO
H
R
N
H
H H
H
H H
O
CH3CN
Et3N
O
O
N
17
HO
HO
MeOH
H2O
THF
O
15
R = OH
16
Na IO4
HO
R =
O
N
OH
H
O
H
O
DCC
DMF
HO
H
H
N
H
H H
O
O
18
Scheme 3: Synthesis of aldehyde derivatives of biotin 14 and deoxycholic acid 18.
As already reported,25 aminooxy function reacts easily with aldehydes and ketones, since such
species could be in the air or glassware (e.g. acetone) or in some solvents (e.g. acetaldehyde).
Hence, it is strongly recommended to avoid a HPLC purification of the crude aminooxy-
oligonucleotides. It is safer to proceed rapidly to the oxime ligation between the crude 3'-
aminooxy-oligonucleotides and the desired aldehydes or ketones to introduce. After ligation,
the conjugates can be easily purified by HPLC. The different 3'-aminooxy-deoxy- and ribo-
oligonucleotides were dissolved in 0.4 M ammonium acetate buffer (pH 4.2) and ketones or
aldehydes were added at rt. After 1 to 4 h of contact time, the conjugates were obtained and
characterized by HPLC and MALDI-TOF MS (See SI). The HPLC profiles showed total
conversion of the starting material and the formation of the conjugate as a single peak, except
for the pyrene conjugate, where the two E and Z isomers were visualized.
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O
1,3
O
N
5
6
O
7
8
O
N
O
3
O
9
3'
O
1,3
O
NH2
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+
AcO-NH4
pH 4.2
O
H
O
1,3
N
O
O
R
O
H
N
H
O
1,3
O
N
R
N
H
O
NH
HN
H
N
O
O
H
O
3
O
S
HN
R=
OH
O
CH3
H
H
HO
CH3
CH3
NH
H
H
H
O
Scheme 4: Conjugation of 3'-aminooxy-(deoxy and ribo)-oligonucleotides with ketone or
aldehyde derivatives.
Table 1: 3-aminooxy-deoxy or ribo-oligonucleotides and their conjugates.
Sequences
MALDI- TOF
calcd found
Deoxy-ribonucleotides
T6-EG3-ONH2
1989.36 1989.79
2029.43 2029.57
2201.61 2201.73
2069.49 2069.51
3814.54 3414.60
3854.60 3854.84
3894.67 3894.65
4026.72 4026.13
5858.80 5858.29
T6-EG3-ON=CMe2
T6-EG3-ON=CH-pyrene
T6-EG3-ON=C(CH2)5
T12-EG3-ONH2
T12-EG3-ON=CMe2
T12-EG3-ON=C(CH2)5
T12-EG3-ON=CH-pyrene
d(TTG TTC GTT GAC CTC CAC T) EG-ONH2
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d(TTG TTC GTT GAC CTC CAC T) EG-ON=CMe2
d(TTG TTC GTT GAC CTC CAC T) EG-ON=CH-fluorene
d(TTG TTC GTT GAC CTC CAC T) EG-ON=CH-dchol
d(TTG TTC GTT GAC CTC CAC T) EG-ON=CH-biot
d(TTG TTC GTT GAC CTC CAC T) EG3-ONH2
d(TTG TTC GTT GAC CTC CAC T) EG3-ON=CMe2
d(TTG TTC GTT GAC CTC CAC T) EG3-ON=CH-fluorene
d(TTG TTC GTT GAC CTC CAC T) EG3-ON=CH-dchol
d(TTG TTC GTT GAC CTC CAC T) EG3-ON=CH-biot
Ribo-oligonucleotides
5898.87 5899.06
6021.00 6019.21
6274.42 6274.21
6126.16 6126.36
5946.91 5947.18
5986.98 5985.32
6109.10 6108.99
6362.53 6361.46
6214.26 6214.29
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r(GUA CCA UGA ACG AU) EG3-ONH2
4679.91 4680.68
4719.88 4719.08
4892.17 4891.40
5095.53 5094.60
4947.26 4946.71
r(GUA CCA UGA ACG AU) EG3-ON=CMe2
r(GUA CCA UGA ACG AU) EG3-ON=CH-pyrene
r(GUA CCA UGA ACG AU) EG3 ON=CH-dchol
r(GUA CCA UGA ACG AU) EG3 ON=CH-biot
CONCLUSION
Solid supports allowing the preparation of oligonucleotides with a 3'-aminooxy function were
rapidly and easily synthesized in few steps from bromoethanol or triethyleglycol. According to
this strategy, it is possible to modulate the length of the spacer between the oligonucleotide and
the aminooxy function since any diol derivatives and ethyleneglycols of any length can be
chosen as a spacer. After elongation by standard phosphoramidite chemistry, a two-step
protocol was applied to first deprotect the oligonucleotides and second to release the 3'-
aminooxy oligonucleotides from the solid support. The corresponding 3'-modified deoxy- and
ribo-oligonucleotides were conjugated to various ketones and aldehydes with high efficiency
by oxime ligation.
This solid supports could be used in combination with other building blocks like special
phosphoramidites to introduce another orthogonal modification at the 5'-end of
oligonucleotides allowing their bis-conjugation. Along this lane, we previously demonstrated
that combination of oxime ligation and CuAAC conjugation are orthogonal.28 The data reported
in this manuscript increases the toolbox to synthesize oligonucleotide conjugates.
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EXPERIMENTAL SECTION
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General Methods: All commercial chemicals were reagent grade and were used without
further purification. The DNA synthesis reagents and phosphoramidites were commercially
available from Link Technologies or ChemGenes. Flash column chromatography was
performed on silica gel 60 (40–63 μm).1H NMR (400 or 500 MHz) and 13C NMR spectra (100
or 126 MHz) were recorded in CDCl3 or DMSO-d6 at room temperature. Compounds 1,36 2a33
and 2b35 were prepared according to literature protocols.
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General protocol of deprotection of aminooxy ethyleneglycol derivatives. To a solution of
phthalimide derivative 2a-b (3.4 mmol) in 12 mL of absolute ethanol, hydrazine hydrate (900
L, 18 mmol) was added. A white precipitate appeared after 5 min and the reaction was stirred
for 3 h. The precipitate was removed by filtration and the solution was evaporated followed by
3 coevaporations with ethanol affording 3a-b that were used without purification.
General protocol for oxime formation. To a solution of aminooxy ethyleneglycol derivatives
3a-b (3.4 mmol) in 10 mL of methanol and 5 mL of DMF, 1 mL of 0.1M ammonium acetate
aqueous buffer, 2 drops of acetic acid and levulinic acid (306 µL, 3.4 mmol) were added. The
solution was stirred at rt for 5 h and the reaction was monitored by RP18 TLC. The solvents
were evaporated. The residue was coevapored 3 times with dry pyridine and compounds 5a-b
were used without purification.
General protocol for dimethoxytritylation. To a solution of compounds 5a-b (3.4 mmol) in
20 mL of dry pyridine, dimethoxytrityl chloride (1.4g, 4.1 mmol) was added. After 2 h under
magnetic stirring, 1 mL of methanol was added. After 5 min, saturated NaHCO3 aqueous
solution (50 mL) was added and the product was extracted with CH2Cl2 (3 × 50 mL). The
organic layers were dried over Na2SO4 and evaporated under vacuum. The residue was purified
by silica gel chromatography from CH2Cl2:Et3N, 98:2, (v/v) to CH2Cl2:Et3N:MeOH 83:2:15,
(v/v/v), to afford compounds 6a-b.
4-((2-dimethoxytrityl-oxyethoxy)imino)-pentanoic acid 6a: Starting from 823 mg (4.0
mmol) of 2a gave 6a 1.15 g 50% (3 steps). 1H-NMR (CDCl3, 400 MHz): δ 1.22 (t, J = 7.3 Hz,
6H, CH3 of Et3N), 1.88 and 1.93 (2s, 3H, CH3), 2.46-2.49 and 2.69-2.73 (2m, 4H,
OCNCH2CH2CO2), 2.99 (q, J = 7.3 Hz, 4H, CH2 of Et3N), 3.24-3.26 (m, 2H, CH2ODMTr),
3.77 (s, 6H, OCH3), 4.17-4.19 (m, 2H, CH2ON=), 6.79-6.82 (m, 4H, Ar), 7.16-7.48 (m, 9H,
13
Ar). C{1H}-NMR (CDCl3, 100 MHz): δ 8.5, 14.4, 19.9, 32.0, 32.6, 44.8, 55.2, 62.7, 72.9,
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85.7, 113.3, 126.9, 128.0, 128.6, 130.3, 136.8, 145.4, 157.1, 158.6, 177.9. HRMS (ESI/Q-TOF):
[M+Na]+ calcd. for C28H31NO6Na 500.2049, found 500. 2053.
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4-((8-dimethoxytrityl-oxy-3,6-dioxaoctanoxy)imino)-pentanoic acid 6b: Starting from 1.0 g
(3.4 mmol) of 2b gave 6b 1.10 g 49% (3 steps). 1H-NMR (CDCl3, 400 MHz): δ 1.23 (t, J = 7.3
Hz, 6H, CH3 of Et3N), 1.83 and 1.87 (2s, 3H, CH3), 2.40-2.63 (m, 4H, CCH2-CH2C), 2.97 (q, J
= 7.3 Hz, 4H, CH2 of Et3N), 3.21-3.24 (m, 2H, CH2ODMTr), 3.66-3.74 (m, 8H, CH2OCH2),
3.78 (s, 6H, OCH3), 4.14-4.17 (t, J = 5.0 Hz, 2H, CH2ON=), 6.80-6.82 (m, 4H, Ar), 7.17-7.47
(m, 9H, Ar). 13C{1H}-NMR (CDCl3, 100 MHz): δ 8.7, 14.5, 20.0, 31.9, 32.3, 44.9, 55.2, 63.2,
69.8, 70.6, 70.7, 70.8, 72.5, 85.9, 113.0, 126.6, 127.7, 128.2, 130.1, 136.2, 145.1, 157.2, 158.6.
HRMS (ESI/Q-TOF): [M + H]+ calcd. for C38H55N2O8 667.3958, found 667.3958.
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General protocol for immobilization on solid support. Compound 6a or 6b (0.46 mmol),
LCAA-CPG (500 mg, 1000 Å), DMAP (56 mg, 0.46 mmol) were dried by coevaporation with
dry pyridine (three times). Dry pyridine (5 mL) was added with EDC (397 mg, 2.1 mmol) and
Et3N (190 L, 1.4 mmol). The mixture was shaken for 24 h at room temperature. The solid
support was filtered off, washed with MeOH and CH2Cl2, and dried. A capping step with
standard Cap A and Cap B solutions was applied for 2 h, and the solid support was filtered off,
washed with MeOH and CH2Cl2, and dried. The trityl assay indicated a loading of 40 mol/g
for 7a and 41 mol/g for 7b.
Synthesis of oligonucleotides 8a-b. The oligonucleotides were elongated from solid support
7a-b (1 μmol scale) on an ABI 394 DNA synthesizer according to standard phosphoramidite
chemistry protocols. Detritylation was performed for 65 s using 3 % trichloroacetic acid (TCA)
in CH2Cl2. For the coupling step: benzylmercaptotetrazole (0.3 M in anhydrous CH3CN) was
used as the activator along with commercially available 2′-deoxyribonucleoside-3′-O-2-
cyanoethyl, N,N-diisopropylphosphoramidites phenoxyacetyl or tert-butylphenoxyacetyl (0.1
M in CH3CN, 30 s coupling time) or 2′-O-pivaloyloximethyl-yribonucleoside-3′-O-2-
cyanoethyl, N,N-diisopropylphosphoramidites (0.1 M in CH3CN, 180 s coupling time). The
capping step was performed using commercially available solutions (Cap A: phenoxyacetic
anhydride:pyridine:THF 10:10:80 v/v/v, and Cap B: 10 % N-methylimidazole in THF) for 60
s. The oxidation step was performed with a standard, diluted iodine solution (0.1 M I2,
THF:pyridine:water 90:5:5, v/v/v) for 15 s.
Deprotection affording 9a-b. For DNA: The CPG was withdrawn in a microtube and treated
with aqueous conc. ammonia for 1 h at rt. The supernatant was removed and CPG was washed
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with water (3 × 1 mL). For RNA: The CPG was treated with a 1 M solution of DBU in dry
acetonitrile for 3 min using two syringes, and washed with dry acetonitrile (3 × 5 mL). Then
the CPG was withdrawn in a microtube and treated with aqueous conc. ammonia for 3 h at 37
°C. The supernatant was removed and CPG was washed with water (3 × 1 mL).
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Release from solid support affording 10a-b. The CPG with the solid-supported
oligonucleotides 9a-b were treated with a solution of 50 mM CH3ONH2,HCl in 0.4 M
ammonium acetate pH 4.2 (2 mL) for 3 h at rt under stirring. The supernatant was applied on a
Sephadex G25 column for desalting. The solution was lyophilized affording the 3'-aminooxy-
(deoxy or ribo)-oligonucleotides 10a-b. Warning: Since aminooxy is a very reactive function,
10a-b should be used rapidly. The glassware must be washed with ethanol to avoided any trace
of acetone.
Conjugation with ketones or aldehyde derivatives. Fluorenone 50 mM in MeOH, Pyrene
carboxyaldehyde 50 mM in DMF, Deoxycholic amide ethanal 50 mM in MeOH, biotin amide
ethanal 50 mM in MeOH. To 3'-aminooxy-EG/EG3-deoxyoligonucleotides 10a-b 15 nmol in
+
25 µL of water were added 225 µM of 0.4M NH4 AcO- pH 4.2 and 10 eq of ketone or aldehyde
derivatives. To 3'-aminooxy-EG3-ribooligonucleotides 10b 5 nmol in 10 µL of water were
+
added 20 µM of 0.4M NH4 AcO- pH 4.2 and 10 eq of ketone or aldehyde derivatives. After 1
h to 4 h the conjugate was purified by size exclusion chromatography on Sephadex G25. The
conversion of 3'-aminooxy-oligonucleotide into its conjugate was better than 90% (See SI).
HPLC purification gave between 19 to 48% of pure conjugate.
Biotin amide 1,2-propanediol 13
To a solution of biotin N-hydroxysuccinimide,17 (150 mg 0.44 mmol) and Et3N (148 µL, 1.06
mmol) in dry CH3CN (10 mL), 3-amino-2-hydroxy-propan-1-ol (48 mg, 0.53 mmol) dissolved
in 3 mL of DMF was added and stirred at rt for 2 h. After evaporation, the residue was purified
by silica gel chromatography from 5% to 20% MeOH in CH2Cl2 affording the diol derivative
1
13, 120 mg, 86%. Rf : 0.5 (MeOH/CH2Cl2, 2:8, v/v). H-NMR (400 MHz, DMSO-d6): ppm
1.23-1.37 (m, 2H, (g) CH2); 1.39-1.66 (m, 4H, (f,h) 2CH2); 2.09 (t, 2H, (e) CH2, J = 7.5 Hz);
2.57 (d, 1H, (l) CH2, J = 12.4 Hz); 2.82 (dd, 1H, (l) CH2, J = 5.2 Hz J = 12.5 Hz); 2.93-3.29 (m,
5H, (a,c) CH2, (i) CH); 3.43-3.49 (m, 1H, (b) CH); 4.1-4.16 (m, 1H, (k) CH); 4.27-4.33 (m, 1H,
(j) CH); 4.51 (t, 1H, OH, J =5.8 Hz); 4.72 (d, 1H, OH, J=4.9 Hz); 6.35 and 6.41 (2s, 2H, (m,n)
13
NH); 7.76 (t, 1H, (d) CONH, J = 5.7 Hz). C{1H}-NMR (100 MHz, DMSO-d6): ppm 25.2,
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27.9, 28.1, 35.0, 39.4, 39.7, 41.9, 55.3, 59.1, 60.9, 63.5, 70.5, 162.6, 172.5. HRMS (ESI/Q-
TOF): [M+H]+ calcd. for C13H24N3O4S, Calcd 318.1482, found 318.1490.
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Biotin amide ethanal 14
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To a solution of biotin amide 1,2-propanediol (120 mg, 0.37 mmol) in CH3OH:H2O, 4:2, (v/v)
(6 mL), sodium periodate (88 mg , 0.41 mmol) was added. After 2 h, the solution was
evaporated and the residue was dissolved in methanol (10 mL). After filtration and evaporation
of filtrate the crude was purified by silica chromatography using a gradient of methanol (5 to
15%) in CH2Cl2 affording 14, 80 mg, 76%. Rf: 0.65 (20% MeOH/CH2Cl2). Mixture of ethanal
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(70%) and dimethylacetal (30%), H-NMR (400 MHz, DMSO-d6): ppm 1.21-1.67 (m, 6H,
(g,f,h), CH2); 2.07 (t, 1.3H, (e) CH2, J = 7.2 Hz); 2.17 (t, 0.7H, (e) CH2, J = 7.2 Hz); 2.57 (d,
1H, (l) CH2, J = 12.5 Hz); 2.82 (dd, 1H, (l) CH2, J = 5.1 Hz J = 12.5 Hz); 3.08-3.13 (m, 1.3H,
(c) CH2 (i) CH); 3.22 (s, 1.8H, OCH3); 3.85 (d, 0.7H, (c) CH2, J = 5.4 Hz); 4.09-4.16 (m, 1H,
(k) CH); 4.27-4.33 (m, 1H, (j) CH); 4.38-4.44 (m, 0.7H, CH(OMe)2); 6.15, 6.18 and 6.35, 6.40
(2 x 2s, 2H, NHCONH); 7.78 and 8.24 (t and bs, 1H, CONH, J = 5.52 Hz); 9.42 (s, 0.7H, CHO).
13C{1H}-NMR (400 MHz, DMSO-d6): ppm 25.2, 27.9, 28.0, 34.9, 39.7, 49.1, 55.3, 59.1, 60.9,
95.7, 162.6, 172.1, 200.1. HRMS (ESI/Q-TOF): [M+H]+ calcd. for C12H20N3O3S, 286.1220,
found 286.1232.
Deoxycholamide 1,2-propanediol 17
To a solution of deoxycholic acid N-hydroxysuccinimide,17 (490 mg, 1.0 mmol) and Et3N (140
µL, 1.1 mmol) in DMF (10 mL), 3-amino-2-hydroxy-propan-1-ol (100 mg, 1.1 mmol) dissolved
in 1 mL of DMF was added and stirred at 50 °C for 3 h. After evaporation the residue was
purified by silica gel chromatography from 5% to 20% of MeOH in CH2Cl2 affording the diol
derivative 17 as a clear oil, 300 mg, 64%. Rf: 0.5 (20% MeOH/CH2Cl2), 1H-NMR (500 MHz,
DMSO-d6): ppm 0.59 (s, 3H, CH3); 0.84 (s, 3H, CH3); 0.91 (d, 3H, CH3, J = 6.5 Hz); 0.95-1.83
(m, 24H, CH2, CH); 1.94-2.14 (m, 2H, C23, CH2); 2.93-3.39 (m, 5H, propyl CH2, CH); 3.45
(qd, 1H, C3, CH, J = 5.4 Hz, J = 10.8 Hz); 3.78 (br, 1H, C12, CH); 4.19 (d, 1H, OH, J = 4.1
Hz); 4.46 (d, 1H, OH, J = 4.3 Hz); 4.50 (t, 1H, OH, J= 5.9 Hz); 4.71 (dd, 1H, OH, J= 1.7 and
4.8 Hz); 7.76 (br, 1H, NH). 13C{1H}-NMR (126 MHz, DMSO-d6): ppm 12.4, 17.0, 23.0, 23.4,
26.0, 26.9, 27.1, 28.5, 30.1, 31.6, 32.3, 32.8, 33.7, 35.0, 35.6, 36.2, 41.5, 41.9, 45.9, 46.1, 47.4,
48.5, 63.5, 69.9, 70.5, 70.9, 173.1. HRMS (ESI/Q-TOF): [M+H]+ calcd. for C27H48NO5, Calcd
466.3532, found 466.3537.
Deoxycholamide ethanal 18
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To a solution of deoxycholamide 1,2-propanediol (280 mg, 0.6 mmol) in CH3OH:H2O:THF,
4:2:2, (v/v/v) (8 mL), sodium periodate (155 mg , 0.73 mmol) was added. After 2 h, the solution
was evaporated and the residue was dissolved in methanol (10 mL). After filtration and
evaporation of filtrate the crude was purified by silica chromatography using a gradient of
methanol (5 to 15%) in CH2Cl2 affording 18 as a clear foam, 240 mg, 92%
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Rf: 0.75 (20% MeOH/CH2Cl2), Mixture of ethanal (20%) and dimethylacetal (80%). 1H-NMR
(400 MHz, DMSO-d6): ppm 0.59 (s, 3H, CH3); 0.84 (s, 3H, CH3); 0.91 (d, 3H, CH3, J = 6.5
Hz); 0.94-1.84 (m, 24H, CH2, CH); 1.92-2.13 (m, 2H, C23, CH2); 3.04 (t, 2H, CH2, J = 5.5 Hz);
3.21 (s, 2H, OMe) 3.34-3.42 (m, 3H, C3 CH, CH2); 3.78 (br, 1H, C12 CH); 4.16 (d, 1H, OH, J
=4.1 Hz); 4.37-4.43 (m, 0.8H, CH(OMe)2); 4.45 (d, 1H, OH, J = 4.3 Hz); 7.76 and 8.22 (2 m,
13
1H, NH); 9.41 (s, 0.2H, CHO). C{1H}-NMR (100 MHz, DMSO-d6): ppm 12.3, 16.9, 23.0,
23.4, 26.0, 26.9, 27.1, 28.5, 30.1, 31.6, 32.2, 32.8, 33.72, 35.0, 35.6, 36.2, 41.5, 43.9, 45.9, 46.1,
47.4, 49.1, 53.4, 54.8, 69.8, 70.9, 95.7, 172.7, 200.1. HRMS (ESI/Q-TOF): [M+H]+ calcd. for
C26H44NO4, 434.3270, found 434.3279.
Associated content
*S Supporting Information. NMR data of 6a-b, 13, 14, 17, 18; HPLC and MALDI-TOF spectra
of 3'-aminooxy-(deoxy or ribo)-oligonucleotides and their conjugates.
AUTHOR INFORMATION
Corresponding Authors
E-mail francois.morvan@umontpellier.fr
ORCID
Dr François Morvan https://orcid.org/0000-0001-5077-0028
Dr Jean-Jacques Vasseur https://orcid.org/0000-0002-4379-6139
Notes
The authors declare no competing financial interest.
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ACKNOWLEDGMENTS
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We thank the Institut des biomolécules Max Mousseron (IBMM) for financial support through
the AAP-IBMM2017 funding program. F. M. is a member of Inserm.
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REFERENCES(1) Juliano, R.; Alam, M. R.; Dixit, V.; Kang, H. Mechanisms and strategies
for effective delivery of antisense and siRNA oligonucleotides. Nucleic Acids Res. 2008, 36,
4158-4171.
(2)
Hill, K. W.; Taunton-Rigby, J.; Carter, J. D.; Kropp, E.; Vagle, K.; Pieken, W.; McGee,
D. P. C.; Husar, G. M.; Leuck, M.; Anziano, D. J.; Sebesta, D. P. Diels-Alder
bioconjugation of diene-modified oligonucleotides. J. Org. Chem. 2001, 66, 5352-5358.
Marchan, V.; Ortega, S.; Pulido, D.; Pedroso, E.; Grandas, A. Diels-Alder
cycloadditions in water for the straightforward preparation of peptide-oligonucleotide
conjugates. Nucleic Acids Res. 2006, 34, e24.
(3)
(4)
(5)
Weisbrod, S. H.; Marx, A. A nucleoside triphosphate for site-specific labelling of DNA
by the Staudinger ligation. Chem. Commun. 2007, 1828-1830.
Bouillon, C.; Meyer, A.; Vidal, S.; Jochum, A.; Chevolot, Y.; Cloarec, J. P.; Praly, J. P.;
Vasseur, J. J.; Morvan, F. Microwave assisted "click" chemistry for the synthesis of
multiple labeled-carbohydrate oligonucleotides on solid support. J. Org. Chem. 2006,
71, 4700-4702.
(6)
(7)
Gramlich, P. M. E.; Wirges, C. T.; Manetto, A.; Carell, T. Postsynthetic DNA
Modification through the Copper-Catalyzed Azide-Alkyne Cycloaddition Reaction.
Angew. Chem. Int. Ed. 2008, 47, 8350-8358.
Pourceau, G.; Meyer, A.; Vasseur, J. J.; Morvan, F. Azide Solid Support for 3 '-
Conjugation of Oligonucleotides and Their Circularization by Click Chemistry. J. Org.
Chem. 2009, 74, 6837-6842.
(8)
(9)
Pourceau, G.; Meyer, A.; Vasseur, J. J.; Morvan, F. Synthesis of Mannose and Galactose
Oligonucleotide Conjugates by Bi-click chemistry. J. Org. Chem. 2009, 74, 1218-1222.
El-Sagheer, A. H.; Brown, T. Click chemistry with DNA. Chem. Soc. Rev. 2010, 39,
1388-1405.
(10) Jewett, J. C.; Bertozzi, C. R. Cu-free click cycloaddition reactions in chemical biology.
Chem. Soc. Rev. 2010, 39, 1272-1279.
(11) Debets, M. F.; Van Berkel, S. S.; Dommerholt, J.; Dirks, A. J.; Rutjes, F.; Van Delft, F.
L. Bioconjugation with Strained Alkenes and Alkynes. Acc. Chem. Res. 2011, 44, 805-
815.
(12) Jadhav, S.; Kakela, M.; Makila, J.; Kiugel, M.; Liljenback, H.; Virta, J.; Poijarvi-Virta,
P.; Laitala-Leinonen, T.; Kyto, V.; Jalkanen, S.; Saraste, A.; Roivainen, A.; Lonnberg,
H.; Virta, P. Synthesis and In Vivo PET Imaging of Hyaluronan Conjugates of
Oligonucleotides. Bioconjugate Chem. 2016, 27, 391-403.
(13) Massaad-Massade, L.; Boutary, S.; Caillaud, M.; Gracia, C.; Parola, B.; Gnaouiya, S.
B.; Stella, B.; Arpicco, S.; Buchy, E.; Desmaele, D.; Couvreur, P.; Urbinati, G. New
Formulation for the Delivery of Oligonucleotides Using "Clickable" siRNA-
Polyisoprenoid-Conjugated Nanoparticles: Application to Cancers Harboring Fusion
Oncogenes. Bioconjugate Chem. 2018, 29, 1961-1972.
(14) Sanchez, A.; Pedroso, E.; Grandas, A. Maleimide-Dimethylfuran exo Adducts:
Effective Maleimide Protection in the Synthesis of Oligonucleotide Conjugates. Org.
Lett. 2011, 13, 4364-4367.
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7
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(15) Sanchez, A.; Pedroso, E.; Grandas, A. Conjugation Reactions Involving Maleimides and
Phosphorothioate Oligonucleotides. Bioconjugate Chem. 2012, 23, 300-307.
(16) Sanchez, A.; Pedroso, E.; Grandas, A. Easy introduction of maleimides at different
positions of oligonucleotide chains for conjugation purposes. Org. Biomol. Chem. 2012,
10, 8478-8483.
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(17) Meyer, A.; Vasseur, J. J.; Morvan, F. Synthesis of Monoconjugated and Multiply
Conjugated Oligonucleotides by "Click Thiol" Thiol-Michael-Type Additions and by
Combination with CuAAC "Click Huisgen". Eur. J. Org. Chem. 2013, 465-473.
(18) Ligeour, C.; Dupin, L.; Marra, A.; Vergoten, G.; Meyer, A.; Dondoni, A.; Souteyrand,
E.; Vasseur, J. J.; Chevolot, Y.; Morvan, F. Synthesis of galactoclusters by metal-free
thiol "click chemistry" and their binding affinity for Pseudomonas aeruginosa lectin
LecA. Eur. J. Org. Chem. 2014, 7621–7630.
(19) Meyer, A.; Noel, M.; Vasseur, J. J.; Morvan, F. Hetero-Click Conjugation of
Oligonucleotides with Glycosides Using Bifunctional Phosphoramidites. Eur. J. Org.
Chem. 2015, 2921-2927.
(20) Knall, A.-C.; Slugovc, C. Inverse electron demand Diels-Alder (iEDDA)-initiated
conjugation: a (high) potential click chemistry scheme. Chem. Soc. Rev. 2013, 42, 5131-
5142.
(21) Busskamp, H.; Batroff, E.; Niederwieser, A.; Abdel-Rahman, O. S.; Winter, R. F.;
Wittmann, V.; Marx, A. Efficient labelling of enzymatically synthesized vinyl-modified
DNA by an inverse-electron-demand Diels-Alder reaction. Chem. Commun. 2014, 50,
10827-10829.
(22) Trevisiol, E.; Renard, A.; Defrancq, E.; Lhomme, J. The oxyamino-aldehyde coupling
reaction - an efficient method for the derivatization of oligonucleotides. Tetrahedron
Lett. 1997, 38, 8687-8690.
(23) Salo, H.; Virta, P.; Hakala, H.; Prakash, T. P.; Kawasaki, A. M.; Manoharan, M.;
Lonnberg, H. Aminooxy functionalized oligonucleotides: Preparation, on-support
derivatization, and postsynthetic attachment to polymer support. Bioconjugate Chem.
1999, 10, 815-823.
(24) Defrancq, E.; Lhomme, J. Use of an aminooxy linker for the functionalization of
oligodeoxyribonucleotides. Bioorg. Med. Chem. Lett. 2001, 11, 931-933.
(25) Forget, D.; Boturyn, D.; Defrancq, E.; Lhomme, J.; Dumy, P. Highly efficient synthesis
of peptide-oligonucleotide conjugates: Chemoselective oxime and thiazolidine
formation. Chem. Eur. J. 2001, 7, 3976-3984.
(26) Katajisto, J.; Virta, P.; Lonnberg, H. Solid-phase synthesis of multiantennary
oligonucleotide glycoconjugates utilizing on-support oximation. Bioconjugate Chem.
2004, 15, 890-896.
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23
24
25
26
27
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44
45
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53
54
55
56
57
58
59
60
(27) Forget, D.; Renaudet, O.; Boturyn, D.; Defrancq, E.; Dumy, P. 3'-Oligonucleotides
conjugation via chemoselective oxime bond formation. Tetrahedron Lett. 2001, 42,
9171-9174.
(28) Meyer, A.; Spinelli, N.; Dumy, P.; Vasseur, J. J.; Morvan, F.; Defrancq, E.
Oligonucleotide Sequential Bis-Conjugation via Click-Oxime and Click-Huisgen
Procedures. J. Org. Chem. 2010, 75, 3927-3930.
(29) Singh, Y.; Murat, P.; Defrancq, E. Recent developments in oligonucleotide conjugation.
Chem. Soc. Rev. 2010, 39, 2054-2070.
(30) Cieslak, J.; Grajkowski, A.; Ausin, C.; Gapeev, A.; Beaucage, S. L. Permanent or
reversible conjugation of 2'-O- or 5'-O-aminooxymethylated nucleosides with
functional groups as a convenient and efficient approach to the modification of RNA
and DNA sequences. Nucleic Acids Res. 2012, 40, 2312-2329.
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(31) Defrancq, E.; Hoang, A.; Vinet, F.; Dumy, P. Oxime bond formation for the covalent
attachment of oligonucleotides on glass support. Bioorg. Med. Chem. Lett. 2003, 13,
2683-2686.
(32) Edupuganti, O. P.; Defrancq, E.; Dumy, P. Head-to-tail oxime cyclization of
oligodeoxynucleotides for the efficient synthesis of circular DNA analogues. J. Org.
Chem. 2003, 68, 8708-8710.
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13
14
15
16
17
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20
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24
25
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27
28
29
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31
32
33
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(33) Lonnberg, T.; Suzuki, Y.; Komiyama, M. Prompt site-selective DNA hydrolysis by
Ce(IV)-EDTA using oligonucleotide multiphosphonate conjugates. Org. Biomol. Chem.
2008, 6, 3580-3587.
(34) Schmidt, P.; Zhou, L. N.; Tishinov, K.; Zimmermann, K.; Gillingham, D. Dialdehydes
Lead to Exceptionally Fast Bioconjugations at Neutral pH by Virtue of a Cyclic
Intermediate. Angew. Chem. Int. Ed. 2014, 53, 10928-10931.
(35) Meyer, A.; Vasseur, J. J.; Dumy, P.; Morvan, F. Phthalimide-Oxy Derivatives for 3-or
5-Conjugation of Oligonucleotides by Oxime Ligation and Circularization of DNA by
"Bis- or Tris-Click" Oxime Ligation. Eur. J. Org. Chem. 2017, 6931-6941.
(36) Angeli, A.; Dupin, L.; Madaoui, M.; Li, M. C.; Vergoten, G.; Wang, S.; Meyer, A.;
Gehin, T.; Vidal, S.; Vasseur, J. J.; Chevolot, Y.; Morvan, F. Glycoclusters with
Additional Functionalities for Binding to the LecA Lectin from Pseudomonas
aeruginosa. Chemistryselect 2017, 2, 10420-10427.
(37) Lavergne, T.; Bertrand, J. R.; Vasseur, J. J.; Debart, F. A Base-Labile Group for 2 '-OH
Protection of Ribonucleosides: A Major Challenge for RNA Synthesis. Chem. Eur. J.
2008, 14, 9135-9138.
(38) Piper, P. W.; Clark, B. F. C. Selective Reaction of Methoxyamine with Cytidine
Residues in Mammalian Initiator Transfer Ribonucleic-Acid. Nucleic Acids Res. 1974,
1, 45-51.
(39) The aldehydes were obtained as a mixture with the dimethylacetals which were
converted in situ into the aldehydes species thanks to the acidic buffer used for the
coupling by oxime ligation
16
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