A new safety-catch protecting group and linker for solid-phase synthesis

Article abstract of DOI:10.1016/j.tetlet.2010.04.047

The use of two derivatives of 2-methoxy-4-methylsulfinylbenzyl alcohol is demonstrated as a safety-catch protecting group and linker for solid-phase peptide synthesis. The protecting group and linker are stable to TFA and are readily removed under reducti

Full text of DOI:10.1016/j.tetlet.2010.04.047

Tetrahedron Letters 51 (2010) 3218–3220
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Tetrahedron Letters
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A new safety-catch protecting group and linker for solid-phase synthesis
*
Sathiah Thennarasu, Chuan-Fa Liu
Division of Chemical Biology and Biotechnology, School of Biological Sciences, Nanyang Technological University, 60 Nanyang Drive, Singapore 637 551, Singapore
a r t i c l e i n f o
a b s t r a c t
Article history:
The use of two derivatives of 2-methoxy-4-methylsulfinylbenzyl alcohol is demonstrated as a safety-
catch protecting group and linker for solid-phase peptide synthesis. The protecting group and linker
are stable to TFA and are readily removed under reductive acidolytic conditions.
Ó 2010 Elsevier Ltd. All rights reserved.
Received 4 February 2010
Revised 30 March 2010
Accepted 12 April 2010
Available online 18 April 2010
Orthogonal protecting groups are widely used in the solution
and solid-phase synthesis of natural products, small molecules
and complex peptides.1 Consequently, various protecting groups
that can be removed under selective conditions have been devel-
oped.2 For instance, the two widely used solid-phase peptide syn-
thesis methods involve, respectively, Fmoc/t-Bu and Boc/Bzl
orthogonal protection strategies.3 Incomplete coupling due to re-
duced solubility of the growing peptide chain and inter- and in-
tra-chain interactions are major drawbacks encountered during
Fmoc/t-Bu-based peptide synthesis.4 These difficulties can be over-
come by employing the Boc/Bzl method and solvents with high sol-
vating parameter such as CH2Cl2, (CF3)2CHOH and TFA.5 In the case
of the Boc/Bzl strategy, the use of acids such as hazardous HF and
trifluoromethanesulfonic acid (TFMSA) precludes the synthesis of
side-chain-protected peptides required for head-to-tail cyclization
and convergent synthesis. Semipermanent protecting groups and
linkers based on the TFA-stable p-methylsulfinylbenzyl ester have
previously been reported.6 Reductive acidolysis conditions have
been employed for removal of the protecting group and cleavage
of the peptide from the resin. However, the Bzl-type protecting
groups are also removed under the reported conditions.7 Selective
removal of Bzl-type protecting groups while the peptide is at-
tached to the resin, and selective cleavage of the peptide ester bond
while the Bzl-type side-chain protecting groups are intact, would
be useful in head-to-tail cyclization and convergent synthesis.
In this Letter, we report the potential of two derivatives of 2-
methoxy-4-methylsulfinylbenzyl alcohol or Mmsb-OH, 7 and 14,
for the protection of amine and carboxylic acid groups, respec-
tively. The protecting groups are structurally related to 4-meth-
ylsulfinylbenzyl alcohol-based Msib8 and Msz9 which were
developed for the protection of hydroxy groups. Since the sulfoxide
moiety para to the hydroxymethyl group is known to impart
exceptional acid-stability to the ester bond,6b,8 we envisioned that
the presence of a methoxy group ortho to the hydroxymethyl group
would render the ester bond more labile, especially during reduc-
tive acidolysis. As a logical extension of this proposition we also
evaluated carbamate-based amine protecting group, which
would be useful for the preparation of N-protected peptide acids
using the Fmoc/t-Bu strategy.
2-Methoxy-4-methylsulfinylbenzyl alcohol 5 was readily pre-
pared in four steps as shown in Scheme 1. Commercially available
3-methoxythiophenol was alkylated using an equimolar quantity
of MeI and a slight excess of triethylamine. Formylation of 2 using
DMF–POCl3 at 50 °C gave 2-methoxy-4-methylthiobenzaldehyde 3
as the major product owing to the strong p-directing ability of the
thioether. The major isomer was isolated using column chromatog-
raphy. Reduction of 3 with sodium triacetoxyborohydride gave the
corresponding alcohol 4 which was subsequently oxidized using
a
HS
O
S
O
S
O
ii)
i)
CHO
1
3
2
iii)
O
S
S
O
O
iv)
OH
OH
4
5
v)
O
O
S
O
S
O
vi)
O
O
O
O
O
O
NH
NH
HO
O
6
7
Scheme 1. Synthesis of Mmsb-OH 5 and Mmsz-Gly-OH. Reagents and conditions:
(i) MeI, Et3N, CH2Cl2, rt, 6 h; (ii) DMF–POCl3, DMF, 50 °C, 15 h, 55% for two steps;
(iii) NaB(OAc)3 , Et2O, 0 °C to rt, 3–5 h, 80%; (iv) mCPBA, CH2Cl2, 0 °C, 2–3 h, 80%; (v)
(a) CDI, DMF, 3 h and (b) H-Gly-OtBu, DMF, 6 h, 48%; (vi) TFA–CH2Cl2 (1:1), rt, 0.5 h,
71%.
* Corresponding author. Tel.: +65 6316 2867; fax: +65 6791 3856.
E-mail address: cfliu@ntu.edu.sg (C.-F. Liu).
0040-4039/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tetlet.2010.04.047

S. Thennarasu, C.-F. Liu / Tetrahedron Letters 51 (2010) 3218–3220
3219
mCPBA to the corresponding sulfoxide 5. The oxidation was carried
out under controlled conditions whereby small portions of wet
mCPBA (50–55%) were added over a period of 2–3 h to a stirred
solution of 4 in ice-cold conditions to give only the sulfoxide prod-
uct 5 in a very good yield.
For the preparation of Mmsz-protected glycine 6, a slight excess
of CDI was reacted with 1 equiv of Mmsb-OH 5 for 3 h and then
with 2 equiv of glycine t-butyl ester. The Mmsz-protected glycine
ester 6 was treated with 50% TFA/CH2Cl2 to liberate the carboxylic
acid group. The overall yield involving all the steps in Scheme 1
was 12%. The NMR and mass spectral data were in agreement with
the proposed structure (see Supplementary data).
In order to establish the suitability of the Mmsz-group for the
safety-catch semipermanent protection of an amine group, a
model hexapeptide corresponding to human Histone H3 (130–
135) was synthesized on Wang resin using the Fmoc/t-Bu strategy
(Scheme 2). Mmsz-Gly-OH was coupled to the resin-bound peptide
using HBTU and DIEA.
The final cleavage was achieved with TFA/H2O (97.5:2.5) (25 °C,
1 h) or TFA/(i-Pr)3SiH/H2O (92.5:5.0:2.5) (25 °C, 6 h). While the for-
mer cleavage conditions gave Mmsz-protected peptide 8 in excel-
lent purity, the latter rendered the completely deprotected peptide
9 (Fig. 1). It is imperative to note that Mmsz protection of the
amine group is stable to TFA but can be deprotected completely
under reductive acidolysis conditions without requiring the use
of a strong acid such as HF or TFMSA. For peptides containing
Trp, Tyr and Cys residues, phenol and mercaptoethanol can be
added as scavengers to obtain Mmsz-protected peptides. Consider-
ing the different acidolytic conditions reported in the literature8
and the simple cleavage mixture used in this study, the Mmsz pro-
tective group appears to be a potential alternative to other benzyl-
oxycarbonyl-based protecting groups, especially in solid-phase
synthesis.
We also explored the possibility of using Mmsb-OH as a safety-
catch linker vis-à-vis a semipermanent protecting group for car-
boxylic acids. A derivative of Mmsb-OH 14 was prepared for this
purpose as shown in Scheme 3, and its utility as a linker and semi-
permanent protecting group for the carboxylic end of a peptide
was demonstrated as shown in Scheme 4. First, ethyl bromoacetate
was attached to the free thiol 1 to give ester 10. Vilsmeier formy-
lation of 10 gave two isomers and the major isomer 11 was isolated
by column chromatography. After reduction of the aldehyde 11 to
12 and oxidation to the corresponding sulfoxide 13, the carboxylic
acid 14 was liberated by saponification using 2 N NaOH and subse-
quent acidification with 2 N HCl. The overall yield was 30%. The
Mmsb-OH derivative 14 was anchored to the commercially
Figure 1. RP HPLC profiles of peptide 8 (trace 1) and peptide 9 (trace 2).
O
S
O
HS
O
i)
O
10
1
ii)
O
O
O
S
O
iii)
S
O
O
O
O
OH
CHO
12
iv)
11
O
S
O
O
v), vi)
OH
O
S
OH
HO
O
13
14
PyBOP, DIEA
O
H2N
O
S
HN
O
Mmsb-AM-Resin
HO
15
Scheme 3. Synthesis of an Mmsb linker for solid-phase synthesis. Reagents and
conditions: (i) BrCH2COOEt, Et3N; (ii) DMF–POCl3; (iii) NaB(OAc)3H; (iv) mCPBA; (v)
2 N NaOH; (vi) 2 N HCl. See Supplementary data for experimental details.
Wang Resin
1. (Fmoc-Ala)2O, cat. DMAP; Fmoc deprotection
2. Fmoc-amino acid coupling/ deprotection cycles
3. Mmsz-Gly-OH, HBTU, DIEA
HO
O
O
S
Mmsb-AM-Resin
HN
O
HO
Mmsz-Gly-Ile-Arg(Pbf)-Gly-Glu(OtBu)-Arg(Pbf)-Ala-O
TFA/H2O
1. (Boc-Ala)2O, cat.DMAP; 33%TFA/CH2Cl2
2. Boc-SPPS cycles
Mmsz-Gly-Ile-Arg-Gly-Glu-Arg-Ala-OH
Boc-Glu(OBn)-Val-Leu-Phe-Ala
Cleavage
Mmsb
8
TFA/TIS/H2O
0 o
C
TFA/DMS/NH4I
H-Gly-Ile-Arg-Gly-Glu-Arg-Ala-OH
H2N-Glu(OBn)-Val-Leu-Phe-Ala-OH
9
16
Scheme 2. Synthesis of a model peptide in Mmsz-protected or deprotected form
under different cleavage conditions.
Scheme 4. Boc solid-phase synthesis of a small model peptide using Mmsb resin.

3220
S. Thennarasu, C.-F. Liu / Tetrahedron Letters 51 (2010) 3218–3220
We envision that the Mmsz-group would be useful as a
semipermanent protecting group for selected amines, while the
Mmsb-ester would serve as a safety-catch carboxylic acid-protect-
ing group which could be exploited as a linker in solid-phase
synthesis of complex molecules that require selective removal of
the side-chain protecting groups during synthesis. This work pro-
vides a new addition to the repertoire of orthogonal protecting
groups that are available for organic synthesis.
Acknowledgements
We thank the Ministry of Education, Singapore, for financial
support as well as Nanyang Technological University. We also
thank Mr. Yang Renliang for help with Fmoc SPPS.
Supplementary data
Supplementary data (details on the synthesis of Mmsz-pro-
tected glycine and the Mmsb-OH derivative (linker), and NMR
and mass spectrometric measurements) associated with this arti-
cle can be found, in the online version, at doi:10.1016/
j.tetlet.2010.04.047.
Figure 2. RP HPLC profile of the TFA/DMS/NH4I-cleaved peptide 16.
available aminomethyl polystyrene resin using PyBOP and DIEA.
The resin was treated with 2% hydrazine in DMF to regenerate
any benzyl alcohol that was acylated in the previous step.
To test the suitability of the Mmsb-OH derivative as a protect-
ing group and linker in peptide synthesis, a model pentapeptide
was synthesized following standard Boc-chemistry protocols.
Boc-Alanine was attached to the linker using the symmetric anhy-
dride of Boc-Alanine and catalytic amounts of DMAP. The Boc-pro-
tecting group was removed using 33% TFA in DCM. Subsequent
amino acids were coupled using HBTU as the coupling reagent.
The presence and absence of free amino groups were monitored
during each step by the Kaiser test, which also indicated the stabil-
ity of the peptide ester bond to repetitive treatment with 30% TFA
in CH2Cl2. The peptide ester bond was finally cleaved from the re-
sin using TFA/DMS/NH4I10 at 0 °C (2 Â 30 min) and the crude prod-
ucts were analyzed by HPLC and ESI-MS. The yield of the crude
peptide after lyophilization was about 64%. The cleavage cocktail
TFA/TIS/H2O was also capable of cleaving the peptide from the
safety-catch linker at room temperature and took 6–7 h for com-
plete cleavage. On the other hand, treatment of the peptidyl resin
with TFA/H2O (9.5:0.5) mixture did not give any detectable
amounts of the peptide. It is clear from Figure 2 that the side-chain
benzyl ester (protecting group) on Glu in peptide 16 remained in-
tact under the conditions used for cleavage of the peptide ester
bond with Mmsb-AM-Resin.
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