Peptide cyclization via ring-closing metathesis: The N-alkenoxy peptide approach

Article abstract of DOI:10.1039/b812611a

The preparation of cyclic hexapeptides from N-hydroxy tripeptides building blocks is described. Introduction of an unsaturated chain on the hydroxamate oxygen followed by fragment coupling leads to N,N′-dialkenoxy hexapeptides that are efficiently cyclize

Full text of DOI:10.1039/b812611a

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PAPER
www.rsc.org/obc | Organic & Biomolecular Chemistry
Peptide cyclization via ring-closing metathesis: the N-alkenoxy
peptide approach†
James Lawrence, Muriel Jourdan, Yannick Valle´e and Ve´ronique Blandin*
Received 25th July 2008, Accepted 9th September 2008
First published as an Advance Article on the web 17th October 2008
DOI: 10.1039/b812611a
The preparation of cyclic hexapeptides from N-hydroxy tripeptides building blocks is described.
Introduction of an unsaturated chain on the hydroxamate oxygen followed by fragment coupling leads
to N,N¢-dialkenoxy hexapeptides that are efficiently cyclized through a ring-closing metathesis reaction.
The length of the alkene chains allows the modulation of the ring size: the synthesis of 17- and
18-membered cycles is reported.
amino acids, O-allyl- serine, homoserine or tyrosine can be used.13
Introduction
Nevertheless, in all cases, side-chains involved in the ring closure
are not available anymore to interact with other molecules, such
as molecular receptors. When biological properties are at stake,
this loss of interaction can modify the response of the peptide
more than the studied cyclization does. Thus, another possibility
is the introduction of an alkene functionality at the nitrogen of
peptide bonds14 but here also the unsaturated amino acid has to
be prepared before its incorporation in the peptide chain.
We recently described the preparation of N-hydroxy peptides
and their modification to N-acyloxy peptides through acylation
of the hydroxamate oxygen.15 It is thus possible to introduce
unsaturated acyl groups in the peptide. However, these acyl
groups are rather reactive and the corresponding cyclized peptide
would probably not be stable in vivo. We therefore considered
that N-alkenoxy peptides (Scheme 1) could present an interesting
alternative for the cyclization of peptides via RCM.
The preparation and study of cyclic analogues is a standard
procedure in the development of therapeutical agents starting
from one identified linear peptide binding to a target receptor.
Constraining peptides through cyclization was first introduced
as a tool to investigate bioactive conformations of hormones.1
These researches led the way to considerable developments in
drug design: cyclization often results in higher receptor binding
affinities than the linear counterpart thanks to a smaller loss of
entropy in the binding; moreover, the constrained structure can
also enhance the selectivity of the ligand and its stability towards
enzymatic degradation.2
With the discovery of air-stable and functional-group tolerant
catalysts, ring-closing olefin metathesis3 (RCM) has emerged as a
powerful tool to prepare cyclic peptides. RCM has for instance
been used in the synthesis of molecules binding to the Grb2 SH2
domain4 or in the development of inhibitors of the hepatitis C virus
NS3 protease.5 Analogues of cystine-6 or lanthionine7-containing
biologically active peptides have been prepared by RCM-mediated
substitution of a carbon-carbon double bond for the disulfide
or monosulfide bridge. Introduction, through RCM, of an all-
hydrocarbon link between two amino acid residues has also been
studied as a way to stabilize b-turn8 or a- and 310-helices.9,10
The cyclization staples the linear peptide between two amino
acid residues referred to as i and i + n positions. The unsatu-
rated chains involved in the ring closure can be introduced via
derivatization of the terminal amine and/or carboxylic acid,11
as part of an amino acid side-chain, or by modification of the
peptide backbone. The most common solution is the incorporation
of an amino acid presenting an unsaturated side-chain such as
allylglycine, homo- and bishomo-allylglycine or more complex
molecules.12 This strategy lacks flexibility since several peptides
have to be prepared if the ring size is to be studied. On the other
hand, to cope with the preparation (or cost) of these unnatural
Scheme 1 RCM-mediated cyclization of N,N¢-dialkenoxy peptide.
In this article, we demonstrate the feasibility of this approach
via the convergent synthesis of N,N¢-dialkenoxy hexapeptides
1 and 2 (Fig. 1) and their RCM-mediated cyclization to form
17- and 18-membered rings.
De´partement de Chimie Mole´culaire, UMR-5250, ICMG FR-2607, CNRS,
Universite´ Joseph Fourier, BP-53, 38041, Grenoble Cedex 9, France. E-mail:
Veronique.Blandin@ujf-grenoble.fr; Fax: +33 476 635 983; Tel: +33 476 514
803
† Electronic supplementary information (ESI) available: Copies of 1H and
13C NMR spectra for all new compounds, representative structure of 11
and 12 from conformational analysis data. See DOI: 10.1039/b812611a
Fig. 1 Target N,N¢-dialkenoxy hexapeptides.
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obtained in good yields by reaction of tripeptides 7a and 9,
and 7b and 10 respectively, using the coupling agent EDCI (1-
ethyl-3-(3¢-dimethylaminopropyl)-carbodiimide hydrochloride) in
conjunction with HOBt (1-hydroxybenzotriazole). Ring-closing
was then performed through a metathesis reaction in the presence
of Grubbs’ catalyst II (2nd generation),19 under high dilution
conditions.20
Results and discussion
A fragment coupling strategy was chosen as it allows the obtention
of a N,N¢-dialkenoxy hexapeptide presenting two alkene chains
of different lengths. The synthesis (Scheme 2) started from the
previously reported reduction of oxime 3.15
Liquid chromatography allowed the separation of the di-
astereomeric N-hydroxy dipeptides 4a and 4b. They were then
coupled with Fmoc-protected alanine acid chloride16 in the
presence of sodium hydrogencarbonate in dichloromethane. In
these conditions selective N-acylation is possible.15,17 O-Alkylation
of the hydroxamate group in the N-hydroxy tripeptides 5a and 5b
was performed by reaction with 3-buten-1-ol under Mitsunobu
conditions,18 leading to the N-homoallyloxy peptides 6a and 6b.
Cleavage of the benzylic ester using BCl3 gave the corresponding
carboxylic acid fragments 7a and 7b. On the other hand, reaction
of the N-hydroxy tripeptide 5a with allyl alcohol under Mitsunobu
conditions allowed the preparation of N-allyloxy peptide 8.
Piperidine-mediated deprotection of the terminal amine of 8
gave the amine fragment 9. Similarly, deprotection of 6b led
to the second amine fragment 10. Hexapeptides 1 and 2 were
The 17-membered cyclic peptide 11 was isolated in 85% yield.
1
According to H and 13C NMR spectra, the cyclization led to
a single stereoisomer. A vicinal coupling constant of 15.4 Hz
could be determined for the ethylenic proton HA, indicating an E
geometry for the alkene bond. Other examples of RCM-mediated
formation of 17-membered rings in peptides involve connections
between allylglycine residues at i and i + 4 positions.21 Using
Grubbs’ catalyst II, mixture of E and Z isomers in various
proportions were obtained. The cyclic peptide 11 is, to our
knowledge, the first example of an (i, i + 3)-linked 17-membered
cyclic peptide prepared through RCM.
On the other hand, several RCM-mediated connections between
residues at i and i + 3 positions leading to 18-membered cycles are
described. They involve bishomoallylglycine22 or O-allylserine10,11b
Scheme 2 Preparation and cyclization of N,N¢-dialkenoxy hexapeptides 1 and 2.
4576 | Org. Biomol. Chem., 2008, 6, 4575–4581 This journal is The Royal Society of Chemistry 2008
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residues. In the latter case, cyclization of a tetrapeptide gave a
1:1 mixture of E and Z isomers.23 Longer peptide sequences
and/or Ca-tetrasubstituted a-amino acid residues – such as 2-
aminoisobutyric acid residues (Aib) – induce conformational
restrictions and thus higher selectivities are obtained: up to 20:1
in favor of the E isomer when an Aib-rich octapeptide is at stake.10
In the case of the hexapeptide 2, the 18-membered ring 12 was
obtained in 71% yield as a mixture of stereoisomers. Analysis
of the spectra from 1H NMR and COSY experiments (in CDCl3)
revealed that several proton resonances were splitted into two: NH
Ala4 (3:1 ratio), CHa Val5, CHa Val3. A vicinal coupling constant
of 15.4 Hz could be determined for one ethylenic proton. From
this estimation, we may conclude that the geometry of the major
isomer is E, in accordance with literature results.
In order to determine some structural features of 11 and
12, further NMR studies and restrained molecular dynamics
using an explicit chloroform solvent model were performed. The
structures obtained from molecular modeling are representative of
the experimental NOE.24 The cyclic parts of molecules 11 and 12
have similar backbone features but do not present any particular
secondary structure. Nevertheless, the presence, in the NOESY
spectra, of significant correlations between the ethylenic protons
and the Ha proton of Ala4 and Val5 indicates the cyclization
constrains the peptide.
Perkin Elmer 341 polarimeter; the corresponding concentration is
1
reported in g per 100cm-3. H NMR (300 MHz) and 13C NMR
(75 MHz) spectra were run on a Bruker Advance300 spectrometer,
and obtained from CDCl3 (dC 77.2 ppm; standard for 1H spectra:
tetramethylsilane dH 0.0 ppm). Multiplicities are reported as
follows: s = singlet, d = doublet, t = triplet, q = quadruplet,
m = multiplet, dd = doublet of doublet, dt = doublet of triplet;
coupling constants J and dA - dB (for AB spin system) are reported
1
in Hz. H and 13C resonance assignments were performed using
conventional 1D and 2D techniques (DEPT, COSY, HMQC and
HMBC experiments). Infrared (IR) spectra were obtained either
from neat films, from a thin film of a dichloromethane solution of
the compound on sodium chloride discs, or from a dispersion
of the compound in a KBr plate. IR spectra were recorded
on a Nicolet Impact-400 FT-IR spectrometer and the data are
reported as absorption maxima in cm-1. Mass spectra (LRMS)
were recorded on a ThermoFinnigan PolarisQ EI/CI ion-trap
spectrometer (DCI; ammonia-isobutane 62:38). High resolution
mass spectra (HRMS) were recorded at the LCOSB, UMR 7613,
Universite´ Pierre et Marie Curie, Paris. Experimental errors for
HRMS data are estimated between 1 and 2 ppm.
Fmoc-Ala1-w[CO-N(OH)]-Val2-Val3-OBn (5b)25. To a solu-
tion of 4b (185.8 mg, 576 mmol) in CH2Cl2 (15 mL) were added
NaHCO3 (60.1 mg, 715 mmol) and Fmoc-Ala-Cl (220.1 mg,
667 mmol). The mixture was stirred overnight. Evaporation of
the solvent under reduced pressure followed by purification by
flash chromatography (Silica gel, CH2Cl2-MeOH 100:0 to 98:2)
gave 5b (148.5 mg, 241 mmol, 42% yield) as a white solid which
can be recrystallized from CH2Cl2/cyclohexane. Mp: 112–113 ◦C.
Conclusions
Alkylation of N-hydroxy peptides building blocks allows the
introduction of alkene chains in the peptide at a late stage of the
synthesis. Modulation of the ring size is thus easy to achieve. These
N-alkenoxy tripeptides can be elongated using a classical peptide
synthesis sequence. In the resulting cyclized peptide, all the native
amino acid residues are still available for molecular recognition.
The N-alkenoxy peptides approach we propose for the RCM-
mediated cyclization of peptides could be a versatile option for the
incorporation of a constraining moiety into biologically relevant
peptides.
25
Rf 0.44 (CH2Cl2-MeOH 95:5). [a]D +19.8 (c 1.50 in CHCl3). dH
(300 MHz; CDCl3) 8.73 (1H, s, NOH), 8.47 (1H, d, J 9.1, NH
Val3), 7.73 (2H, d, J 7.6, CHar Fmoc), 7.67–7.60 (2H, m, CHar
Fmoc), 7.45-7.15 (9H, m, CHar), 6.54 (1H, d, J 9.1, NH Ala1),
5.30-5.20 (1H, m, CHa Val2), 5.16 (2H, ABq, J 12.0, dA-dB 56.2,
CH2 -OBn), 5.13-5.02 (1H, m, CHa Ala1), 4.74 (1H, dd, J 4.5
and 9.2, CHa Val3), 4.42-4.15 (3H, m, CH Fmoc, CH2 Fmoc),
2.50-2.33 (1H, m, CHiPr Val2), 2.25-2.12 (1H, m, CHiPr Val3), 1.29
(3H, d, J 6.9, CH3 Ala1), 0.99 (6H, d, J 6.7, CH3 Val2), 0.84 (3H,
d, J 6.9, CH3 Val3), 0.73 (3H, d, J 6.9, CH3 Val3). dC (75 MHz;
Experimental (see also ESI†)
=
=
=
CDCl3) 173.2 (C O Val2), 172.6 (C O Ala1), 171.7 (C O Val3),
=
156.2 (C O Fmoc), 144.2, 144.1, 141.40, 141.37 (Car Fmoc), 135.1
General experimental methods
(Car -OBn), 128.8, 128.7, 127.8, 127.1, 125.5, 125.4, 120.0 (CHar),
67.6, 67.1 (CH2 -OBn, CH2 Fmoc), 63.3 (CHa Val2), 56.6 (CHa
Val3), 47.4 (CH Fmoc), 47.1 (CHa Ala1), 32.0 (CHiPr Val3), 27.6
(CHiPr Val2), 19.9, 19.1, 18.9, 18.1, 17.5 (CH3). IR: 3378, 3299,
3067, 2964, 2929, 2875, 1724, 1634. HRMS (ESI +) m/z: found,
638.2836; C35H41N3O7Na requires 638.2842.
All non-aqueous reactions were performed under an atmosphere
of argon. Standard inert atmosphere techniques were used in
handling all air and moisture sensitive reagents. CH2Cl2 and
THF were freshly distilled from CaH2 and sodium benzophenone
ketyl respectively. Purchased reagents were used without prior
purification. For chromatographic purification, reagent grade
solvents were used as received. Reactions were monitored by thin
layer chromatography (TLC) using commercial aluminum-backed
silica gel plates (Merck, Kieselgel 60 F254). TLC spots were viewed
under ultraviolet light and by heating the plate after treatment
with an appropriate staining solution (KMnO4, ninhydrin, basic
TTC (2,3,5-triphenyltetrazolium chloride)). Product purification
by flash chromatography was performed using Macherey Nagel
Silica Gel 60M (230–400 mesh).
=
Fmoc-Ala1-w[CO-N(O(CH2)2CH CH2)]-D-Val2-Val3-OBn (6a).
A mixture of 5a (100.0 mg, 162 mmol) and PPh3 (106.4 mg,
406 mmol) was dissolved in THF (10 mL). 3-buten-1-ol (34.7 mL,
406 mmol) and DMAD (methyl azodicarboxylate, 61.4 mL,
408 mmol) were added to the solution. After 4 hours of reaction,
the solvent was evaporated and the crude material was taken
up with CH2Cl2. The mixture was washed with brine and the
aqueous phase was extracted with CH2Cl2. The organic layers
were gathered and dried over Na2SO4, filtrated and evaporated.
Purification by flash chromatography (Silica gel, pentane-EtOAc
Melting points were determined with a Bu¨chi B-540 apparatus
and are given uncorrected. Optical rotations were measured on a
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9:1 to 1:1) gave 6a (47.4 mg, 71 mmol, 44% yield) as a colourless oil.
Rf 0.65 (CH2Cl2-MeOH 95:5). [a]D25 +25.3◦ (c 0.75 in CHCl3). dH
(300 MHz; CDCl3) 7.68 (2H, d, J 7.4, CHar Fmoc), 7.58-7.48 (2H,
m, CHar Fmoc), 7.35-7.15 (9H, m, CHar), 6.95 (1H, d, J 8.2, NH
4.80-4.65 (1H, m, CHa Ala1), 4.42 (1H, dd, J 4.6 and 8.2, CHa
Val3), 4.38-4.31 (2H, m, CHH Fmoc, CH Fmoc), 4.25-4.16 (1H,
m, CHH Fmoc), 4.15-4.02 (2H, m, N–O–CH2), 3.95 (1H, d, J
11.1, CHa Val2), 2.72-2.57 (1H, m, CHiPr Val2), 2.45-2.33 (2H,
m, N–O–CH2–CH2), 2.30-2.15 (1H, m, CHiPr Val3), 1.39 (3H, d,
J 6.8, CH3 Ala1), 1.05-0.80 (12H, m, CH3 Val2, CH3 Val3). dC
=
Val3), 5.80-5.62 (1H, m, CH ), 5.55 (1H, d, J 8.2, NH Ala1), 5.12-
=
4.92 (4H, m, CH2 –OBn, CH2 ), 4.70-4.60 (1H, m, CHa Ala1),
= =
(75 MHz; CDCl3) 174.9, 173.9, 170.6 (C O), 156.0 (C O Fmoc),
4.41 (1H, dd, J 4.6 and 8.4, CHa Val3), 4.33-4.25 (2H, m, CHH
Fmoc, CH Fmoc), 4.22-4.05 (3H, m, CHH Fmoc, N–O–CHH,
CHa Val2), 3.85-3.75 (1H, m, N–O–CHH), 2.55-2.40 (1H, m, CHiPr
Val2), 2.31-2.20 (2H, m, N–O–CH2–CH2), 2.18-2.05 (1H, m, CHiPr
Val3), 1.31 (3H, d, J 6.7, CH3 Ala1), 0.93 (3H, d, J 6.5, CH3 Val2),
0.89 (3H, d, J 6.4, CH3 Val2), 0.83 (3H, d, J 6.9, CH3 Val3), 0.77
(3H, d, J 6.8, CH3 Val3). dC (75 MHz; CDCl3) 174.4, 171.2, 170.3
=
144.1, 144.0, 141.5 (Car Fmoc), 133.5 (CH ), 127.9, 127.3, 125.4,
=
125.3, 120.2 (CHar), 118.2 (CH2 ), 75.9 (N–O–CH2), 74.2 (CHa
Val2), 67.3 (CH2 Fmoc), 57.7 (CHa Val3), 48.1 (CHa Ala1), 47.3
(CH Fmoc), 32.6 (N–O–CH2–CH2), 30.4 (CHiPr Val3), 27.4 (CHiPr
Val2), 19.8, 19.7, 19.4, 18.4, 17.7 (CH3). IR: 3314, 3068, 2965, 2931,
1716, 1651. LRMS (DCI +) m/z: 580.1 (M + H)+, 597.6 (M +
NH4)+. HRMS (ESI +) m/z: found, 618.25684; C32H41N3O7K
requires 618.25761.
=
=
(C O), 155.8 (C O Fmoc), 144.2, 144.0, 141.5, 141.4 (Car Fmoc),
=
135.6 (Car –OBn), 133.7 (CH ), 128.7, 128.5, 128.4, 127.8, 127.2,
=
125.4, 125.3, 120.1 (CHar), 117.9 (CH2 ), 75.9 (N–O–CH2), 68.0
=
Fmoc-Ala1-w[CO-N(O(CH2)2CH CH2)]-Val2-Val3-OH (7b).
(CHa Val2), 67.1, 67.0 (CH2 Fmoc, CH2 -OBn), 57.5 (CHa Val3),
48.0 (CHa Ala1), 47.4 (CH Fmoc), 32.6 (N–O–CH2–CH2), 30.8
(CHiPr Val3), 26.4 (CHiPr Val2), 19.9, 19.7, 19.3, 18.8, 17.7 (CH3).
IR: 3325, 3067, 3028, 2964, 2929, 2868, 1723, 1681, 1649. LRMS
(DCI +) m/z: 687.0 (M + NH4)+. HRMS (ESI +) m/z: found,
708.30366; C39H47N3O7K requires 708.30456.
To a solution of 6b (42.1 mg, 62.9 mmol) in CH2Cl2 (4 mL)
was added BCl3 (1M in hexane) (315 mL, 315 mmol). The
mixture was stirred for 2 hours before a second addition of BCl3
(157 mL, 157 mmol). The mixture was stirred further for 2 hours.
Evaporation of the solvent under reduced pressure followed by
purification by flash chromatography (Silica gel, CH2Cl2-MeOH
98:2 to 90:10) gave 7b (29.2 mg, 50.4 mmol, 80% yield) as a yellow
oil. Rf 0.13 (CH2Cl2-MeOH 95:5). [a]D25 +10.9◦ (c 1.27 in CHCl3).
dH (300 MHz; CDCl3) 7.74 (2H, d, J 7.4, CHar Fmoc), 7.64 (1H,
d, J 7.8, NH Val3), 7.60-7.55 (2H, m, CHar Fmoc), 7.42-7.25 (4H,
=
Fmoc-Ala1-w[CO-N(O(CH2)2CH CH2)]-Val2-Val3-OBn (6b).
The title compound was prepared as described for 6a using 5b
(144.1 mg, 234 mmol). Purification by flash chromatography (Silica
gel, pentane-EtOAc 9:1 to 1:1) gave 6b (74.3 mg, 111 mmol, 47%
25
=
yield) as a colourless oil. Rf 0.56 (CH2Cl2-MeOH 95:5). [a]D
m, CHar Fmoc), 5.85-5.72 (1H, m, CH ), 5.63 (1H, d, J 8.1, NH
Ala1), 5.17-5.05 (2H, m, CH2 ), 4.81-4.69 (1H, m, CHa Ala1),
+4.1◦ (c 1.09 in CHCl3). dH (300 MHz; CDCl3) 7.75 (2H, d, J 7.6,
=
CHar Fmoc), 7.72-7.65 (1H, m, NH Val3), 7.64-7.58 (2H, m, CHar
4.44 (1H, dd, J 4.5 and 8.1, CHa Val3), 4.40-4.30 (2H, m, CHH
Fmoc, CH Fmoc), 4.25-4.15 (1H, m, CHH Fmoc), 4.15-3.95 (2H,
m, N–O–CH2), 4.01 (1H, d, J 11.3, CHa Val2), 2.70-2.55 (1H, m,
CHiPr Val2), 2.43-2.30 (2H, m, N–O–CH2–CH2), 2.29-2.14 (1H, m,
CHiPr Val3), 1.38 (3H, d, J 6.8, CH3 Ala1), 1.02-0.85 (12H, m, CH3
=
Fmoc), 7.43-7.23 (9H, m, CHar), 5.85-5.70 (1H, m, CH ), 5.58 (1H,
=
d, J 7.7, NH Ala1), 5.20-5.00 (4H, m, CH2 –OBn, CH2 ), 4.80-4.70
(1H, m, CHa Ala1), 4.49 (1H, dd, J 4.3 and 8.4, CHa Val3), 4.45-
4.32 (2H, m, CHH Fmoc, CH Fmoc), 4.28-4.20 (1H, m, CHH
Fmoc), 4.10-3.85 (3H, m, N–O–CH2, CHa Val2), 2.75-2.60 (1H,
m, CHiPr Val2), 2.40-2.30 (2H, m, N–O–CH2–CH2), 2.28-2.15 (1H,
m, CHiPr Val3), 1.40 (3H, d, J 6.8, CH3 Ala1), 1.00 (3H, d, J 6.8,
CH3 Val2), 0.97 (3H, d, J 6.8, CH3 Val2), 0.92 (3H, d, J 6.9, CH3
Val3), 0.85 (3H, d, J 6.9, CH3 Val3). dC (75 MHz; CDCl3) 174.6,
=
Val2, CH3 Val3). dC (75 MHz; CDCl3) 174.9, 174.3, 170.4 (C O),
=
=
156.1 (C O Fmoc), 144.1, 144.0, 141.5 (Car Fmoc), 133.5 (CH ),
=
127.9, 127.3, 125.4, 125.3, 120.1 (CHar), 118.2 (CH2 ), 76.0
(N–O–CH2), 73.7 (CHa Val2), 67.3 (CH2 Fmoc), 57.6 (CHa Val3),
48.1 (CHa Ala1), 47.3 (CH Fmoc), 32.6 (N–O–CH2–CH2), 30.5
(CHiPr Val3), 27.5 (CHiPr Val2), 19.8, 19.7, 19.3, 18.4, 17.7 (CH3).
IR: 3319, 3067, 2965, 2932, 2868, 1723, 1652. HRMS (ESI +)
m/z: found, 618.25679; C32H41N3O7K requires 618.25761.
=
=
171.4, 170.3 (C O), 155.8 (C O Fmoc), 144.2, 144.0, 141.5, 141.4
=
(Car Fmoc), 135.7 (Car –OBn), 133.5 (CH ), 128.7, 128.5, 128.4,
=
127.9, 125.4, 125.3, 120.1 (CHar), 118.1 (CH2 ), 75.8 (N–O–CH2),
74.6 (CHa Val2), 67.2, 67.0 (CH2 Fmoc, CH2 -OBn), 57.5 (CHa
Val3), 48.1 (CHa Ala1), 47.4 (CH Fmoc), 32.5 (N–O–CH2–CH2),
30.8 (CHiPr Val3), 27.3 (CHiPr Val2), 19.9, 19.7, 19.3, 18.7, 17.6
(CH3). IR: 3323, 3067, 3033, 2964, 2930, 2868, 2851, 1725, 1681.
HRMS (ESI +) m/z: found, 708.30371; C39H47N3O7K requires
708.30456.
=
Fmoc-Ala1-w[CO-N(OCH2CH CH2)]-D-Val2-Val3-OBn (8).
A mixture of 5a (100.0 mg, 162 mmol) and PPh3 (106.4 mg,
406 mmol) was dissolved in THF (10 mL). Allyl alcohol (28 mL,
412 mmol) and DMAD (methyl azodicarboxylate, 61 mL,
405 mmol) were added to the solution. After 4 hours, the solvent
was evaporated and the crude material was taken up with CH2Cl2.
The mixture was washed with brine and the aqueous phase
was extracted with CH2Cl2. The organic layers were gathered
and dried over Na2SO4, filtrated and evaporated. Purification
by flash chromatography (Silica gel, pentane-EtOAc 9:1 to 1:1)
gave 8 (70.1 mg, 107 mmol, 66% yield) as a colourless oil. Rf
=
Fmoc-Ala1-w[CO-N(O(CH2)2CH CH2)]-D-Val2-Val3-OH (7a).
To a solution of 6a (35.4 mg, 52.9 mmol) in CH2Cl2 (4 mL) was
added BCl3 (1M in hexane) (0.5 mL, 0.5 mmol). The mixture
was stirred overnight. Evaporation of the solvent under reduced
pressure followed by purification by flash chromatography (Silica
gel, CH2Cl2-MeOH 100:0 to 95:5) gave 7a (12.3 mg, 21.2 mmol,
0.56 (CH2Cl2-MeOH 95:5). [a]D +17.7◦ (c 0.83 in CHCl3). dH
25
25
40% yield) as a yellow oil. Rf 0.11 (CH2Cl2-MeOH 95:5). [a]D
(300 MHz; CDCl3) 7.75 (2H, d, J 7.4, CHar Fmoc), 7.63-7.55
(2H, m, CHar Fmoc), 7.43-7.20 (9H, m, CHar), 7.00 (1H, d, J
8.1, NH Val3), 6.00-5.80 (1H, m, CH ), 5.60 (1H, d, J 7.9, NH
Ala1), 5.40-5.25 (2H, m, CH2 ), 5.08 (2H, ABq, J 12.3, dA-dB
+13.0◦ (c 0.71 in CHCl3). dH (300 MHz; CDCl3) 7.75 (2H, d, J
7.6, CHar Fmoc), 7.73-7.65 (1H, m, NH Val3), 7.62-7.53 (2H, m,
CHar Fmoc), 7.42-7.25 (4H, m, CHar Fmoc), 5.85-5.70 (1H, m,
=
=
=
=
CH ), 5.57 (1H, d, J 8.2, NH Ala1), 5.20-5.05 (2H, m, CH2 ),
12.5, CH2 -OBn), 4.85-4.70 (1H, m, CHa Ala1), 4.65-4.55 (1H, m,
4578 | Org. Biomol. Chem., 2008, 6, 4575–4581
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N–O–CHH), 4.49 (1H, dd, J 4.5 and 8.3, CHa Val3), 4.42-4.32
(3H, m, CH Fmoc, CHH Fmoc and N–O–CHH), 4.30-4.20 (2H,
m, CHH Fmoc and CHa Val2), 2.63-2.48 (1H, m, CHiPr Val2),
2.27-2.14 (1H, m, CHiPr Val3), 1.40 (3H, d, J 6.8, CH3 Ala1), 1.00
(3H, d, J 6.9, CH3 Val), 0.97 (3H, d, J 6.9, CH3 Val), 0.91 (3H,
d, J 6.9, CH3 Val), 0.85 (3H, d, J 6.9, CH3 Val). dC (75 MHz;
2929, 2874, 1739, 1682. HRMS (ESI +) m/z: found, 448.28003;
C24H38N3O5 requires 448.28060.
=
Fmoc-Ala1 -W[CO-N(O(CH2)2CH CH2)]-D-Val2 -Val3 -Ala4-
=
W[CO-N(OCH2CH CH2)]-D-Val5-Val6-OBn (1). To a mixture
of 7a (9.8 mg, 16.9 mmol), 9 (8.3 mg, 19.2 mmol) and HOBt (1-
hydroxybenzotriazole, 3.5 mg, 25 mmol) in CH2Cl2 (1.5 mL), was
added EDCI (1-ethyl-3-(3¢-dimethylaminopropyl)-carbodiimide
hydrochloride, 5.0 mg, 26.1 mmol). The mixture was stirred
overnight. The solution was then washed with brine and the
organic layer dried over Na2SO4. Evaporation of the solvent under
reduced pressure followed by purification by flash chromatography
(Silica gel, pentane-EtOAc 9:1 to 1:1) gave 1 (12.0 mg, 12.1 mmol,
=
=
=
CDCl3) 174.5 (C O Ala1), 171.2 (C O Val3), 170.1 (C O Val2),
=
155.8 (C O, Fmoc), 144.2, 144.0, 141.51, 141.46 (CHar Fmoc),
=
135.6 (Car -OBn), 130.8 (CH ), 128.7, 128.53, 128.49, 127.9,
=
127.2, 125.4, 125.3 (CHar), 121.0 (CH2 ), 120.2 (CHar), 77.8
(N–O–CH2), 68.1, 67.2, 67.0 (CH2 –OBn, CH2 Fmoc, CHa Val2),
57.6 (CHa Val3), 48.0 (CHa Ala1), 47.4 (CH Fmoc), 30.9 (CHiPr
Val3), 26.4 (CHiPr Val2), 19.8, 19.7, 19.3, 18.9, 17.7 (CH3 Ala1,
Val2, Val3). IR: 3325, 3065, 2963, 2928, 2874, 1722, 1682, 1655.
LMRS (DCI) m/z: 672.8 (M + NH4)+. HRMS (ESI +) m/z:
found, 694.28843; C38H45N3O7K requires 694.28891.
25
71% yield) as a yellow oil. Rf 0.40 (CH2Cl2-MeOH 95:5). [a]D
+22.8◦ (c 0.69 in CHCl3). dH (300 MHz; CDCl3) 7.76 (2H, d,
J 7.4, CHar Fmoc), 7.65-7.55 (2H, m, CHar Fmoc), 7.45-7.25
(10H, m, CHar, NH Val 6), 7.05-6.90 (1H, m, NH Val3), 6.70-
=
6.60 (1H, m, NH Ala4), 5.95-5.70 (3H, m, 2x CH , NH Ala1),
=
H-Ala1-w[CO-N(OCH2CH CH2)]-D-Val2-Val3-OBn (9). To a
=
5.40-4.95 (6H, m, CH2 -OBn, 2x CH2 ), 4.95-4.86 (1H, m, CHa
Ala4), 4.82-4.75 (1H, m, CHa Ala1), 4.67-4.55 (1H, m, NO–CHH–
solution of 8 (54.0 mg, 82.4 mmol) in THF (4 mL) was added
piperidine (1 mL). The mixture was stirred for 1h. Evaporation
of the solvent under reduced pressure followed by purification
by flash chromatography (Silica gel, CH2Cl2-MeOH 100:0 to
95:5) gave 9 (22.3 mg, 51.4 mmol, 62% yield) as a yellow oil.
=
=
CH ), 4.52-4.35 (3H, m, CHa Val6, CHa Val3, NO–CHH–CH ),
4.34-4.15 (6H, m, CHa Val2, CHa Val5, CH Fmoc, CH2 Fmoc,
N–O–CHH), 4.00-3.90 (1H, m, N–O–CHH), 2.63-2.41 (2H, m,
CHiPr Val2, CHiPr Val5), 2.40-2.29 (2H, m, N–O–CH2–CH2), 2.28-
2.17 (2H, m, CHiPr Val3, CHiPr Val6), 1.45-1.20 (6H, m, CH3 Ala1,
CH3 Ala4), 1.05-0.80 (24H, m, CH3 Val2, CH3 Val5, CH3 Val3, CH3
Val6). dC (75 MHz; CDCl3) 174.2, 171.4, 171.2, 170.1, 169.8 (C O),
155.8 (C O Fmoc), 144.1, 144.0, 141.5 (CH Fmoc), 135.7 (Car
OBn), 133.7 (CH ), 130.9 (NO–CH2–CH ), 128.8, 128.7, 128.54,
128.49, 127.9, 127.3, 125.3 (CHar), 121.0 (N–O–CH2–CH CH2),
Rf 0.11 (CH2Cl2-MeOH 95:5). [a]D +48.3◦ (c 0.64 in CHCl3).
25
dH (300 MHz; CDCl3) 7.45-7.30 (5H, m, CHar), 7.17 (1H, d, J
=
=
8.5, NH Val3), 5.95-5.78 (1H, m, CH ), 5.39-5.23 (2H, m, CH2 ),
5.14 (2H, ABq, J 12.3, dA-dB 14.1, CH2 -OBn), 4.61-4.48 (2H,
m, CHa Val3, N–O–CHH), 4.33-4.17 (2H, m, CHa Val2, N–O–
CHH), 3.85 (1H, q, J 6.8, CHa Ala1), 2.62-2.47 (1H, m, CHiPr
Val2), 2.30-2.16 (1H, m, CHiPr Val3), 1.28 (3H, d, J 6.8, CH3 Ala1),
0.99 (3H, d, J 6.7, CH3 Val2), 0.95 (3H, d, J 6.7, CH3 Val3),
0.93 (3H, d, J 6.9, CH3 Val2), 0.87 (3H, d, J 6.9, CH3 Val3). dC
=
=
–
=
=
=
=
120.2 (CHar), 117.9 (N–O–(CH2)2CH CH2), 77.9, 76.0 (N–O–
CH2), 68.4, 67.9 (CHa Val2, CHa Val5), 67.2, 67.0 (CH2 –OBn,
CH2 Fmoc), 58.9, 57.6 (CHa Val3, CHa Val6), 48.4 (CHa Ala1 or 4),
47.4 (CH Fmoc), 46.6 (CHa Ala4 or 1), 32.7 (N–O–CH2–CH2), 30.8,
30.5 (CHiPr Val3, CHiPr Val6), 26.59, 26.49 (CHiPr Val2, CHiPr Val5),
19.9, 19.8, 19.7, 19.5, 19.2, 18.7, 18.6, 17.9, 17.8, 17.7 (CH3). IR:
3324, 3067, 2965, 2932, 2874, 1724, 1650. HRMS (ESI +) m/z:
found, 1033.50339; C55H74N6O11K requires 1033.50472.
=
(75 MHz; CDCl3) 178.5, 171.6, 170.7 (C O), 135.6 (Car), 131.0
=
=
(CH ), 128.73, 128.69, 128.6, 128.4 (CHar), 120.6 (CH2 ), 77.6
(N–O–CH2), 68.1 (CHa Val2), 67.1 (CH2 -OBn), 57.5 (CHa Val3),
47.8 (CHa Ala1), 30.7 (CHiPr Val3), 26.1 (CHiPr Val2), 20.9, 19.8,
19.7, 19.3, 17.7 (CH3). IR: 3310, 2965, 2929, 2872, 1739, 1685,
1653. LRMS (DCI +) m/z: 434.1 (M + H)+. HRMS (ESI +) m/z:
found, 434.26451; C23H36N3O5 requires 434.26495.
=
Fmoc-Ala1-w[CO-N(O(CH2)2CH CH2)]-Val2-Val3-Ala4-w[CO-
=
N(O(CH2)2CH CH2)]-Val5-Val6-OBn (2). The title compound
=
H-Ala1-w[CO-N(O(CH2)2CH CH2)]-Val2-Val3-OBn (10). To
was prepared as described for 1 using 7b (25.4 mg, 43.8 mmol) and
10 (18.4 mg, 41.1 mmol). Purification by flash chromatography
(Silica gel, pentane-EtOAc 9:1 to 1:1) gave 2 (26.9 mg, 26.7 mmol,
a solution of 6b (47.1 mg, 70.4 mmol) in THF (5 mL) was added
piperidine (1 mL). The mixture was stirred for 1h. Evaporation
of the solvent under reduced pressure followed by purification by
flash chromatography (Silica gel, CH2Cl2-MeOH 100:0 to 90:10)
gave 10 (20.3 mg, 45.4 mmol, 65% yield) as a yellow oil. Rf
25
65% yield) as a yellow oil. Rf 0.40 (CH2Cl2-MeOH 95:5). [a]D
-12.7◦ (c 0.94 in CHCl3). dH (300 MHz; CDCl3) 7.76 (2H, d, J
7.4, CHar Fmoc), 7.63-7.55 (2H, m, CHar Fmoc), 7.47 (1H, d, J
8.7, NH Val6), 7.44-7.26 (10H, m, CHar Fmoc, NH Val3), 6.69
0.16 (CH2Cl2-MeOH 90:10). [a]D +0.8◦ (c 0.92 in CHCl3). dH
25
=
(300 MHz; CDCl3) 7.81 (1H, d, J 8.6, NH Val3), 7.40-7.25 (5H,
(1H, d, J 7.9, NH Ala4), 5.90-5.60 (3H, m, 2x CH , NH Ala1),
=
=
m, CHar), 5.85-5.65 (1H, m, CH ), 5.22-5.05 (4H, m, CH2 -OBn,
5.24-5.02 (6H, m, CH2 -OBn, 2x CH2 ), 5.01-4.90 (1H, m, CHa
=
CH2 ), 4.51 (1H, dd, J 4.5 and 8.8, CHa Val3), 4.02-3.70 (4H, m,
Ala4), 4.83-4.70 (1H, m, CHa Ala1), 4.48 (1H, dd, J 4.3 and 8.4,
CHa Val6), 4.40-3.90 (10H, m, CHa Val3, CHa Val2, CHa Val5, CH
Fmoc, CH2 Fmoc, (2x) N–O–CH2), 2.70-2.50 (2H, m, CHiPr Val2,
CHiPr Val5), 2.50-2.27 (4H, m, (2x) N–O–CH2–CH2), 2.27-2.12
(2H, m, CHiPr Val3, CHiPr Val6), 1.41 (3H, d, J 6.7, CH3 Ala1),
1.32 (3H, d, J 6.9, CH3 Ala4), 1.07-0.80 (24H, m, CH3 Val2, CH3
Val5, CH3 Val3, CH3 Val6). dC (75 MHz; CDCl3) 174.8, 174.2,
171.4, 170.1, 170.0, 169.8 (C O), 155.8 (C O Fmoc), 144.1,
144.0, 141.5 (Car Fmoc), 135.7 (Car –OBn), 133.6, 133.5 (CH ),
128.7, 128.5, 128.4, 127.9, 127.2, 125.3, 120.1 (CHar), 118.1, 118.0
N–O–CH2, CHa Val2, CHa Ala1), 2.75-2.60 (1H, m, CHiPr Val2),
2.40-2.18 (3H, m, N–O–CH2–CH2, CHiPr Val3), 1.29 (3H, d, J
6.8, CH3 Ala1), 0.99 (3H, d, J 6.8, CH3 Val2), 0.95 (3H, d, J 6.8,
CH3 Val3), 0.94 (3H, d, J 6.8, CH3 Val2), 0.88 (3H, d, J 6.8, CH3
=
Val3). dC (75 MHz; CDCl3) 178.5, 171.6, 171.0 (C O), 135.7 (Car),
=
=
133.5 (CH ), 128.7, 128.6, 128.4 (CHar), 118.0 (CH2 ), 75.4 (N–
O–CH2), 74.5 (CHa Val2), 67.0 (CH2 -OBn), 57.5 (CHa Val3), 47.8
(CHa Ala1), 32.5 (N–O–CH2–CH2), 30.8 (CHiPr Val3), 27.2 (CHiPr
Val2), 20.6, 19.9, 19.7, 19.4, 17.6 (CH3). IR: 3308, 3066, 3034, 2964,
=
=
=
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=
(CH2 ), 76.0, 75.9 (N–O–CH2), 73.5, 72.8 (CHa Val2, CHa Val5),
12.3, dA-dB 57.1, CH2 -OBn), 4.85-4.65 (2H, m, CHa Ala1, CHa
Ala4), 4.45 (1H, dd, J 4.4 and 8.3, CHa Val6), 4.40-4.25 (1H, m,
CHH Fmoc), 4.25-4.17 (2H, m, CHH Fmoc, CH Fmoc), 4.15-
4.00 (minor diastereomer: m, CHa Val5, CHa Val3), 3.95-3.75 (m,
4H, NO–CH2 + major diastereomer: m, CHa Val5, CHa Val3),
3.69 (1H, d, J 11.0, CHa Val2), 2.80-2.50 (2H, m, CHiPr Val2,
CHiPr Val5), 2.40-2.00 (4H, m, N–O–CH2–CH2, CHiPr Val3, CHiPr
Val6), 1.50-1.25 (6H, m, CH3 Ala1, CH3 Ala4), 1.10-0.78 (24H, m,
CH3 Val2, CH3 Val3, CH3 Val5, CH3 Val6). dC (75 MHz; CDCl3)
for the major diastereomer: 174.45, 174.38, 172.7, 171.2, 170.4,
67.2, 67.0 (CH2 Fmoc, CH2 –OBn), 58.8 (CHa Val6), 57.4 (CHa
Val3), 48.0 (CHa Ala1), 47.3 (CH Fmoc), 46.1 (CHa Ala4), 32.6,
32.5 (N–O–CH2–CH2), 30.7 (CHiPr Val3, CHiPr Val6), 27.32, 27.28
(CHiPr Val2, CHiPr Val5), 19.9, 19.8, 19.7, 19.5, 19.3, 18.5, 17.9,
17.7 (CH3). IR: 3312, 3067, 2963, 2931, 2874, 1744, 1724, 1682,
1641. HRMS (ESI +) m/z: found, 1047.51887; C56H76N6O11K
requires 1047.52037.
=
Fmoc-Ala1-w[CO-N(O(CH2)2CH *)]-D-Val2-Val3-Ala4-w[CO-
N(OCH2CH* )]-D-Val5-Val6-OBn (11). To a solution of 1
=
=
=
170.2 (C O), 155.8 (C O Fmoc), 144.3, 143.9, 141.5, 141.4 (Car
(7.4 mg, 7.4 mmol) in CH2Cl2 (15 mL), Grubbs’ catalyst II (0.6 mg,
0.7 mmol) was added and the reaction was stirred for 12 hours.
The mixture was washed with brine and extracted with CH2Cl2.
The organic phases were gathered, dried over Na2SO4, filtrated
and evaporated. Flash chromatography (Silica gel, diethyl ether)
gave 11 (6.1 mg, 6.3 mmol, 85% yield) as a colourless oil. Rf
=
Fmoc), 136.0 (Car –OBn), 130.7 (CH ), 128.6, 128.4, 127.8, 127.2
=
(CHar), 126.7 (CH ), 125.4, 125.3, 120.1 (CHar), (overlapped by
CDCl3: N–O–CH2, CHa Val2 or 5), 73.8 (CHa Val2 or 5), 67.2, 66.7
(CH2 –OBn, CH2 Fmoc), 60.8 (CHa Val3), 57.6 (Val6), 48.6 (CHa
Ala1 or 4), 47.4 (CH Fmoc), 46.6 (CHa Ala1 or 4), 31.1, 30.6, 30.3
(2x N–O–CH2–CH2, CHiPr Val3, CHiPr Val6), 27.3 (CHiPr Val2 and 5),
20.5, 19.85, 19.82, 19.7, 19.4, 19.3, 19.2, 19.0, 17.6 (CH3). IR:
3318, 3063, 3033, 2965, 2924, 2872, 1714, 1664. HRMS (ESI +)
m/z: found, 981.53157; C54H73N6O11 requires 981.53318.
0.23 (CH2Cl2-MeOH 95:5). [a]D +26.7◦ (c 0.56 in CHCl3). dH
25
(300 MHz; CDCl3) 7.76 (2H, d, J 7.5, CHar Fmoc), 7.71 (1H, d, J
7.6, NH Val6), 7.63-7.54 (2H, m, CHar Fmoc), 7.44-7.36 (2H, m,
CHar Fmoc), 7.35-7.25 (7H, m, CHar), 6.62 (1H, d, J 6.9, NH Val3),
6.42-6.37 (1H, m, NH Ala4), 5.87-5.72 (2H, m, N–O–CH2–CHB=,
Analysis of conformational preferences of 11 and 12
=
NH Ala1), 5.57 (1H, dt, J 6.5 and 15.4, CHA ), 5.14 (2H, ABq,
NMR experiments were carried out using a Varian Unity+
500 MHz spectrometer. The NOESY experiments were recorded
with 512 (t1) ¥ 2048 (t2) complex data points and 64 scans were
acquired over a spectra width of 5000 Hz in both dimensions.
They were acquired with mixing times of 250 and 400 ms. Data
processing and analysis were performed using the VNMR program
(Varian) on a Sunray workstation.
J 12.5, dA-dB 8.3, CH2 -OBn), 4.92-4.81 (1H, m, CHa Ala1), 4.78-
4.70 (1H, m, CHa Ala4), 4.70-4.60 (1H, m, N–O–CHH–CH ), 4.57
(1H, d, J 11.0, CHa Val2), 4.50 (1H, dd, J 5.7 and 7.9, CHa Val6),
4.42-4.22 (4H, m, CH2 Fmoc, CH Fmoc, CHa Val5), 4.18-4.08
=
=
(1H, m, N–O–CHH–CH ), 4.05-3.84 (3H, m, N–O–CH2–CH2,
CHa Val3), 2.59-2.44 (2H, m, CHiPr Val2, CHiPr Val5), 2.42-2.28
(1H, m, N–O–CH2–CHH), 2.25-2.12 (1H, m, N–O–CH2–CHH),
2.08-1.96 (2H, m, CHiPr Val3, CHiPr Val6), 1.45-1.40 (3H, m, CH3
Ala1), 1.27-1.20 (3H, m, CH3 Ala4), 1.09-0.80 (24H, m, CH3 Val2,
CH3 Val3, CH3 Val5, CH3 Val6). dC (75 MHz; CDCl3) 175.3, 175.1,
Molecular modeling: calculations were performed on a worksta-
tion using InsightII 2005 and Discover 2004.1 (Biosym/Molecular
Simulation). The molecules 11 and 12 were constructed in InsightII
using the Biopolymer module. In the case of 12, we only studied
the major E isomer. Calculations were performed using an explicit
=
=
171.9, 170.6, 170.2, 169.3 (C O), 156.0 (C O Fmoc), 144.2, 143.9,
=
141.53, 141.46 (Car Fmoc), 135.8 (Car –OBn), 135.7 (CH ), 128.7,
˚
solvent model in chloroform (cubic box of 40 A side length and
=
128.5, 128.3, 127.94, 127.91, 127.2 (CHar), 125.6 (CH ), 125.4,
417 CHCl3 molecules) and the cvff force-field. Periodic bounded
conditions were applied.
125.2, 120.2 (CHar), 76.5, 75.3 (N–O–CH2), 67.4, 66.9 (CH2 –OBn,
CH2 Fmoc, CHa Val2 or 5), 64.5 (CHa Val2 or 5), 61.2, 58.0 (CHa Val3,
CHa Val6), 48.2, 48.1 (CHa Ala1 and 4), 47.3 (CH Fmoc), 34.4 (N–
O–CH2–CH2), 30.8, 30.5 (CHiPr Val3, CHiPr Val6), 27.6, 26.8 (CHiPr
Val2, CHiPr Val5), 20.3, 19.95, 19.85, 19.5, 19.4, 19.3, 18.8, 18.1,
17.5 (CH3). IR: 3320, 2961, 2925, 2872, 2853, 1736, 1660. HRMS
(ESI +) m/z: found, 1005.47242; C53H70N6O11K requires
1005.47342.
After minimization (100 steps of steepest descents then 2000 of
conjugate gradients), a restrained molecular dynamics protocol
was applied. The structure was heated to 900 K in 1.5 ps, then
slowly cooled to 300 K by steps of 100 K. At each step the system
was allowed to stay 10 ps. 100 structures were saved over the last
10 ps dynamics calculation and each was subjected to a further
minimization (2000 steps of conjugate gradient). The 6 structures
with the lowest energy were averaged and energy minimized to
lead to the representative structure.
=
Fmoc-Ala1 -W[CO-N(O(CH2)2CH *)]-Val2 -Val3 -Ala4 -W[CO-
=
N(O(CH2)2CH* )]-Val5-Val6-OBn (12). To a solution of 2
(18.8 mg, 18.6 mmol) in CH2Cl2 (15 mL), Grubbs’ catalyst II
was added in three times: 1.6 mg (1.9 mmol); 0.8 mg (0.9 mmol)
after 6h; 0.8 mg (0.9 mmol) after 12h. The reaction was stirred
further for 2 hours. The mixture was washed with brine and
extracted with CH2Cl2. The organic phases were gathered, dried
over Na2SO4, filtrated and evaporated. Flash chromatography
(Silica gel, pentane-EtOAc 9:1 to 1:1) gave 12 (12.9 mg, 13.2 mmol,
71% yield) as a 3:1 mixture of two stereoisomers. Rf 0.27 (CH2Cl2-
MeOH 95:5). dH (300 MHz; CDCl3) 7.75 (2H, d, J 7.4, CHar
Fmoc), 7.63-7.50 (2H, m, CHar Fmoc), 7.46 (1H, d, J 7.0, NH
Val6), 7.42-7.25 (10H, m, CHar, NH Val3), 6.37-6.28 and 6.00-
The restraints used in the calculations were derived from
observed intermolecular NOE classified semiquantitatively into
˚
˚
three categories, strong (1.8–3.0 A), medium (2.5–4.0 A) and weak
˚
˚
˚
(3.5–5.0 A). Only one violation greater than 0.2 A (0.25 A) was
detected in the averaged structures.
Acknowledgements
The authors thank Beatrice Gennaro for recording some 2D
NMR spectra, Marie-Louise Dheu-Andries for her help with the
chloroform box for the dynamics calculation and Pierre Yves
Chavant for useful discussions during the preparation of this
manuscript. We are grateful to Universite´ Joseph Fourier and
=
5.95 (1H, ratio 3:1, m, NH Ala4), 5.75-5.55 (2H, m, CH , NH
=
Ala1), 5.55-5.43 (1H, dt, J 6.1 and 15.4, CH ), 5.12 (2H, ABq, J
4580 | Org. Biomol. Chem., 2008, 6, 4575–4581
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©

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the CNRS for financial support and to the CECIC for providing
computer facilities.
1879–1880 and the recent Account: (c) A. Patgiri, A. L. Jochim and
P. S. Arora, Acc. Chem. Res., 2008, DOI: 10.1021/ar700264k and ref.
therein.
10 A. K. Boal, I. Guryanov, A. Moretto, M. Crisma, E. L. Lanni, C.
Toniolo, R. H. Grubbs and D. J. O’Leary, J. Am. Chem. Soc., 2007,
129, 6986–6987.
11 (a) See in particular ref. 4b, 9b, c; (b) See also: U. Kazmaier, C. Hebach,
A. Watzke, S. Maier, H. Mues and V. Huch, Org. Biomol. Chem., 2005,
3, 136–145.
Notes and references
1 See in particular: (a) V. J. Hruby, Life Sci., 1982, 31, 189–199and
references therein; (b) H. Kessler, Angew. Chem. Int. Ed. Engl., 1982,
21, 512–523and ref. therein.
2 For a review, see: (a) P. Li and P. P. Roller, Curr. Top. Med. Chem.,
2002, 2, 325–341and ref. therein; (b) For a recent study of the effect of
backbone cyclization, see: S. Hess, Y. Linde, O. Ovadia, E. Safrai, D. E.
Shalev, A. Swed, E. Halbfinger, T. Lapidot, I. Winkler, Y. Gabinet, A.
Faier, D. Yarden, Z. Xiang, F. P. Portillo, C. Haskell-Luevano, C. Gilon
and A. Hoffman, J. Med. Chem., 2008, 51, 1026–1034.
3 See for instance the review: (a) D. Astruc, New J. Chem., 2005, 29,
42–56, and the Nobel lectures: (b) Y. Chauvin, Angew. Chem. Int. Ed.,
2006, 45, 3741–3747; (c) R. R. Schrock, Angew. Chem. Int. Ed., 2006,
45, 3748–3759; (d) R. H. Grubbs, Angew. Chem. Int. Ed., 2006, 45,
3760–3765.
12 (a) See ref. 4–8, 9a. See also: (b) P. W. R. Harris and M. A. Brimble,
Org. Biomol. Chem., 2006, 4, 2696–2709 (c) N. Schmiedeberg and H.
Kessler, Org. Lett., 2002, 4, 59–62; (d) A. S. Ripka, R. S. Bohacek
and D. H. Rich, Bioorg. Med. Chem. Lett., 1998, 8, 357–360; (e) J.
Pernerstorfer, M. Schuster and S. Blechert, Chem. Commun., 1997,
1949–1950.
13 (a) See ref. 5a, 6e, 10, 11b. See also: (b) H. E. Blackwell, J. D. Sadowsky,
R. J. Howard, J. N. Sampson, J. A. Chao, W. E. Steinmetz, D. J. O’Leary
and R. H. Grubbs, J. Org. Chem., 2001, 66, 5291–5302; (c) S. J. Miller,
H. E. Blackwell and R. H. Grubbs, J. Am. Chem. Soc., 1996, 118,
9606–9614.
4 (a) F. Liu, K. M. Worthy, L. K. Bindu, R. J. Fisher and T. R. Burke, Jr.,
J. Org. Chem., 2007, 72, 9635–9642 and ref. therein; (b) F. J. Dekker,
N. J. de Mol, M. J. E. Fischer, J. Kemmink and R. M. J. Liskamp, Org.
Biomol. Chem., 2003, 1, 3297–3303.
14 (a) See ref. 8c, 9b, 12b, 13c. See also: (b) J. Cluzeau, S. Oishi, H. Ohno,
Z. Wang, B. Evans, S. C. Peiper and N. Fujii, Org. Biomol. Chem., 2007,
5, 1915–1923; (c) J. F. Reichwein, C. Versluis and R. M. J. Liskamp,
J. Org. Chem., 2000, 65, 6187–6195 and ref. therein (d) J. F. Reichwein,
B. Wels, J. A. W. Kruijtzer, C. Versluis and R. M. J. Liskamp, Angew.
Chem. Int. Ed., 1999, 38, 3684–3687.
5 (a) F. Vela´zquez, S. Venkatraman, W. Wu, M. Blackman, A. Prongay,
V. Girijavallabhan, N.-Y. Shih and F. G. Njoroge, Org. Lett., 2007, 9,
3061–3064; (b) Y. S. Tsantrizos, J.-M. Ferland, A. McClory, M. Poirier,
V. Farina, N. K. Yee, X. Wang, N. Haddad, X. Wei, J. Xu and L. Zhang,
J. Organomet. Chem., 2006, 691, 5163–5171 and ref. therein.
6 See for instance: (a) D. D’Addona, A. Carotenuto, E. Novellino, V.
Piccand, J. C. Reubi, A. Di Cianni, F. Gori, A. M. Papini and M.
Ginanneschi, J. Med. Chem., 2008, 51, 512–520; (b) A. J. Robinson, J.
Elaridi, B. J. Van Lierop, S. Mujcinovic and W. R. Jackson, J. Pept.
Sci., 2007, 13, 280–285; (c) A. Mollica, G. Guardiani, P. Davis, S.-W.
Ma, F. Porreca, J. Lai, L. Mannina, A. P. Sobolev and V. J. Hruby,
J. Med. Chem., 2007, 50, 3138–3142; (d) I. Berezowska, N. N. Chung,
C. Lemieux, B. C. Wilkes and P. W. Schiller, J. Med. Chem., 2007,
50, 1414–1417; (e) J. L. Stymiest, B. F. Mitchell, S. Wong and J. C.
Vederas, J. Org. Chem., 2005, 70, 7799–7809; (f) V. Chaleix, V. Sol, M.
Guilloton, R. Granet and P. Krausz, Tetrahedron Lett., 2004, 45, 5295–
5299.
7 (a) V. R. Pattabiraman, J. L. Stymiest, D. J. Derksen, N. I. Martin and
J. C. Vederas, Org. Lett., 2007, 9, 699–702and ref. therein; (b) N. Ghalit,
J. F. Reichwein, H. W. Hilbers, E. Breukink, D. T. S. Rijkers and R. M. J.
Liskamp, ChemBioChem, 2007, 8, 1540–1554and ref. therein.
8 b-turn mimics via RCM-mediated cyclization between i and i + 3 amino
acid residues: (a) J. Giovannoni, C. Didierjean, P. Durand, M. Marraud,
A. Aubry, P. Renaut, J. Martinez and M. Amblard, Org. Lett., 2004, 6,
3449–3452; (b) S. Hanessian and M. Angiolini, Chem. Eur. J., 2002, 8,
111–117. Between i and i + 1: see for instance: (c) J. Einsiedel, H. Lanig,
R. Waibel and P. Gmeiner, J. Org. Chem., 2007, 72, 9102–9113and ref.
therein.
15 J. Lawrence, L. Cointeaux, P. Maire, Y. Valle´e and V. Blandin, Org.
Biomol. Chem., 2006, 4, 3125–3141.
16 Many amino acids, including amino acids bearing a functionalized side
chain, can be introduced via the Fmoc-protected amino acid chloride.
see: (a) L. A. Carpino, B. J. Cohen, K. E. Stephens, Jr., S. Y. Sadat-
Aalaee, J.-H. Tien and D. C. Langridge, J. Org. Chem., 1986, 51,
3732–3734, and for instance a recent example using Fmoc-Ser(t-Bu)-Cl:
(b) A. C. Spivey, J. McKendrick, R. Srikaran and B. A. Helm, J. Org.
Chem., 2003, 68, 1843–1851.
17 P. Maire, V. Blandin, M. Lopez and Y. Valle´e, Synlett, 2003, 671–674.
18 The Mitsunobu reaction occurs under mild conditions and can be
carried out in the presence of a wide range of functional groups. See
the recent review: (a) T. Y. S. But and P. H. Toy, Chem. Asian. J., 2007, 2,
1340–1355and ref. therein. The reaction is compatible with usual amino
acid side chain protections. See for instance the study of N-methylation
of Na-arylsulfonyl peptides via Mitsunobu reaction: (b) E. Biron, J.
Chatterjee and H. Kessler, J. Peptide Sci., 2006, 12, 213–219.
19 M. Scholl, S. Ding, C. W. Lee and R. H. Grubbs, Org. Lett., 1999, 1,
953–956.
20 Concentrations: 0.5–1.2 mM. Byproducts that could result from
intermolecular RCM or cross metathesis reactions were not recovered
during the purification process.
21 (a) See ref. 6f, 7a. See also: (b) N. Ghalit, D. T. S. Rijkers and R. M. J.
Liskamp, J. Mol. Catal. A, 2006, 254, 68–77.
22 See ref. 8b; the stereoselectivity of the ring-closing reaction is not given.
23 This result was obtained using the first generation of Grubbs’ catalyst
(ref.11b).
9 (a) F. Bernal, A. F. Tyler, S. J. Korsmeyer, L. D. Walensky and G. L.
Verdine, J. Am. Chem. Soc., 2007, 129, 2456–2457 and ref. therein;
(b) D. Wang, M. Liu and P. S. Arora, Angew. Chem. Int. Ed., 2008, 47,
24 See Experimental and Electronic Supplementary Information.
25 Compounds 3, 4a, 4b and 5a were previously described (ref.15).
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