Azopeptides: Synthesis and Pericyclic Chemistry

Article abstract of DOI:10.1021/acs.orglett.5b02723

Azopeptides possess an imino urea as an amino amide surrogate in the sequence. Azopeptides were synthesized by oxidation of aza-glycine residues and employed in pericyclic chemistry. Diels-Alder cyclizations and Alder-ene reactions on azopeptides enabled construction of constrained aza-pipecolyl and reactive aza-allylglycyl residues. X-ray crystallographic analyses of azopeptide 16a and azapeptides 30a and 35a provided insight into imino urea configuration and conformational affects of cycloalkane side chains at the semicarbazide α- and β-nitrogen, respectively.

Full text of DOI:10.1021/acs.orglett.5b02723

Letter
pubs.acs.org/OrgLett
Azopeptides: Synthesis and Pericyclic Chemistry
Ramesh Chingle and William D. Lubell*
́ ́ ́ ́ ́
Departement de Chimie, Universite de Montreal, C.P. 6128, Succursale Centre-Ville, Montreal, Quebec Canada H3C 3J7
S
* Supporting Information
ABSTRACT: Azopeptides possess an imino urea as an amino amide surrogate in the sequence. Azopeptides were synthesized by
oxidation of aza-glycine residues and employed in pericyclic chemistry. Diels−Alder cyclizations and Alder−ene reactions on
azopeptides enabled construction of constrained aza-pipecolyl and reactive aza-allylglycyl residues. X-ray crystallographic analyses
of azopeptide 16a and azapeptides 30a and 35a provided insight into imino urea configuration and conformational affects of
cycloalkane side chains at the semicarbazide α- and β-nitrogen, respectively.
n the study of biologically active molecules, conformational
constraint provides fundamental insight for designing enzyme
dynamics of side-chain and backbone conformation.13 To open
potential for pericyclic chemistry in peptide frameworks, we have
now created azopeptides 5 and studied their Diels−Alder and
Alder−ene chemistry to synthesize aza-pipecolyl and aza-
allylglycine residues.
I
inhibitors and receptor modulators.1 In particular, the application
of semicarbazides as amino acid surrogates in azapeptides (e.g., 1,
Figure 1) can restrict backbone geometry to enhance selectivity
Aza-glycine analogues 6−12 were synthesized using protocols
featuring acylation of the peptide chain with activated
methylidene or alkyl carbazates prior to deprotection and chain
elongation (see the Supporting Information).14−16 Azopeptides
were then produced by oxidation of the aza-glycine residues using
N-bromosuccinimide (NBS, Scheme 1) and pyridine in CH2Cl2
at −78 °C to room temperature.3d,17
Carbamates 8−12 were converted, respectively, to azopeptides
13−17 using the NBS/pyridine conditions, as indicated by the
change of the reaction solution color to pale yellow and
observation of a new relatively nonpolar bright yellow spot on
the TLC plate. Suitably pure azopeptides for subsequent
chemistry were isolated in 97−99% yields after concentration
of the reaction mixture, partitioning between aqueous sodium
bicarbonate and ethyl acetate, and evaporation of the organic
phase. In the IR spectra, azopeptides exhibited a band between
1700 to 1765 cm−1 indicative of the NN stretching vibration.18
Azopeptides 13−17 were used without further purification
because they were relatively unstable and decomposed over time,
as observed by the appearance of additional spots on TLC.
Certain azopeptides (e.g., 14 and 16) could be studied by NMR
spectroscopy. Moreover, crystals of sarcosine analogue 16a were
isolated and characterized by X-ray diffraction (Figure 2).
A structure search of the Cambridge Crystallographic Data
Center found only ethyl (N-phenylcarbamoyl)azoformate (18)
as a simple diacyl diazine to compare with 16a.19 Azopeptide 16a
and azoformate 18 exhibited similar NN bond lengths (∼1.24
Figure 1. Aza- and azopeptides and aza-pipecolyl analogues.
and stability.2 Azapeptides bearing cyclic and unsaturated
residues are consequently important targets, respectively,
because of their enhanced conformational rigidity and notable
reactivity.3,4 For example, aza-pipecolyl peptidomimetics (e.g.,
2−4) have displayed antibacterial and antidiabetic activities
because of their inhibitory activity on peptide deformylase and
dipeptidyl peptidase IV.3b,5,6
The introduction of cyclic and unsaturated aza-residues is,
however, challenging, due to the need to selectively differentiate
neighboring nitrogens, and accomplished typically by lengthy
hydrazine chemistry using ionic intermediates.2a In contrast,
pericyclic chemistry offers atom-economical access to cyclic and
unsaturated systems. Azopeptides 5 possessing NN bonds
offer potential for pericyclic reactions with diene and ene systems,
which have typically been performed using simple azodicarbox-
ylates,7 carbamoyl diazine carboxylates,8 N-alkyltriazole diones,9
and phthalazinedione.10,11 Azo-bridges between aromatic pep-
tide side chains have been made12 and used to photoregulate
Received: September 22, 2015
© XXXX American Chemical Society
A
DOI: 10.1021/acs.orglett.5b02723
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Azopeptides 13−17 were examined as dienophiles in Diels−
Alder reactions with butadiene, 2,3-dimethylbutadiene, cyclo-
pentadiene, and cyclohexadiene. Butadiene was used in a sealed
tube. Azopeptides 13−17 reacted smoothly with the butadienes
and cyclopentadiene in dichloromethane at −78 °C to rt over 24
h to give the aza-pipecolyl analogues 19−29 in 92−99% yields
after purification by silica gel chromatography (Scheme 2). 2-
Scheme 1. Synthesis of Azopeptides 13−17
Scheme 2. Aza-pipecolyl Peptide Synthesis from Diels−Alder
Reactions of Azopeptides 13−17
Figure 2. Crystal structure of azopeptide 16a with bond lengths and
dihedral angles (C = gray, H = green, N = blue, O = red).
Methyl-1,3-butadiene reacted with azopeptide 13a to give a 1:1
regioisomeric mixture of Diels−Alder adducts 21a in 92% yield
(see the SI). Cycloadditions with cyclopentadiene gave 1:1−4:1
diastereomeric mixtures measured using super critical fluid
chromatography (SFC). The NMR spectra of Diels−Alder
adducts from butadiene and 2,3-dimethylbutadiene were
recorded at 100−120 °C to resolve broad signals due to tertiary
urea isomers. Molecular ions characteristic of Diels−Alder
adducts were observed from reactions between cyclohexadiene
and azopeptide (e.g., 13a);however, their NMRspectra exhibited
an additional set of downfield vinyl protons indicative of Alder−
ene reaction products (Scheme 3; see the SI).20
Alder−ene reactions were examined with two different
conditions using azopeptides 13a and 15a−16a and three
olefins: 1,3-cyclohexadiene, cycloheptatriene, and isobutylene.
Aza-allylglycines 30a−32a and 34a were, respectively, obtained
using cyclohexadiene and isobutylene in CH2Cl2 at −78 °C to rt
overnight. Isobutylene was used in a sealed tube. Under these
conditions, no product was isolated from the reaction of
cycloheptatriene and azopeptide 13a. In contrast, cyclohepta-
triene and isobutylene reacted, respectively, with azopeptide 13a
in the presence of NBS and pyridine at −78 °C in CH2Cl2 to give
β-substituted analogues 35a and 33a in 28% and 52% yield. Aza-
glycine 8a was isolated with 35a by chromatography of the
reaction mixture of azopeptide 13a, likely due to loss of
cycloheptatrienyl cation and protonation on silica gel. The
Alder−ene reaction typically provided α-substituted product
under both conditions A and B (e.g., 30a and 34a); however, in
cases where no reaction was observed with conditions A, β-
substituted product was obtained using conditions B.
Å) and E-diazine configuration; the former was slightly distorted
from planarity with a dihedral angle value of −173°. Ureas 16a
and 18 differed in their respective E and Z conformations.
Torsion angles for 16a and 18 were, respectively, 20° and −177°,
indicative of greater twisting in the tertiary urea. Azopeptide 16a
adopted a bent conformation; however, in spite of their relative
proximity, no seven-, eight-, or 10-membered hydrogen bonds
were observed between the amide NH group and either diacyl
diazine nitrogen or carbonyl groups. In the crystal lattice,
azopeptides 16a stacked antiparallel to each other forming
intermolecular hydrogen bonds between the benzhydrylamide
NH proton and sarcosine carbonyl oxygen.
In CD3OD, the NMR spectrum of 16a exhibited a 1:1 isomeric
mixture. Isomer assignment to the diazine or tertiary urea was not
possible, but evidence for isomerization of diazine 15a was
obtained by NMR spectroscopy, which indicated a 1:3 mixture of
azo-isomers in CDCl3.
Switching the N-terminal residue from carbamate to amide in
aza-glycyl peptides 6 and 7 possessing alanyl and phenylacetyl
residues, respectively, destabilized the oxidation product.
Although the characteristic azopeptide yellow spot was observed
by TLC analysis after treatment of 6b with NBS at −78 °C for 10
min, the color was pale and other products were present.
Attempts to trap the diazine with butadiene and isobutylene
produced complex mixtures observed using LCMS, albeit minor
peaks corresponded to the masses of the Diels−Alder and Alder−
ene adducts. Attempts failed to make azopeptide using
(diacetoxyiodo)benzene and lead tetraacetate to oxidize aza-
glycines 6 and 7.
B
DOI: 10.1021/acs.orglett.5b02723
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Scheme 3. Aza-allylglycine Synthesis by Alder−Ene Reactions
Figure 4. X-ray structures of 30a and 35a: broken lines represent
inferred hydrogen bonds.
Table 1. Structures and ϕ and ψ Dihedral Angles (in degrees)
from Crystal Analyses and Ideal Turns
Alder−ene regiochemistry was determined by a combination
of NMR spectroscopy, MS experiments, and X-ray analyses (see
the SI). Azapeptides 33a and 34a were examined by
heteronuclear multiple-bond correlation (HMBC) NMR spec-
troscopy using nonuniform sampling at 120 °C, which resolved
the proton signals at distinct chemical shifts (Figure 3; see the SI).
entry
type I β-turn22b
ϕi+1
ψi+1
ϕi+2
ψi+2
−60
−30
−90
0
30a
−60.9
−58.1
−60
−33.0
−24.7
120
−79.0
−66.7
−90
−14.2
−17.7
0
Boc-azaAla-Pro-NHiPr (38)21a
type VIa β-turn22b
35a
−65.0
153.7
−81.9
−17.8
−169.7°. The ϕ and ψ dihedral angles of the X-ray structures for
30a and 35a are compared in Table 1 with ideal turn geometry
and the crystal structure of Boc-azaAla-Pro-NHiPr 38, which also
adopted a type-I β-turn geometry.21,22 Aza-residue substituent
position influenced turn geometry, offering intriguing potential as
a design element for controlling peptide and peptoid
conformation.
Figure 3. HMBC correlations used to assign the regiochemistry of the
ene adducts 33a and 34a.
The practical utility of this method was demonstrated by the
synthesis of aza-pipecolyl analogues of melanocyte-stimulating
hormone release inhibiting factor-1 (MIF-1, Pro-Leu-Gly-
NH2).23 A bioactive β-turn conformation has been proposed
for the positive allosteric modulator activity of this endogenous
neuropeptide on the D2 and D4 dopamine receptor
subtypes,24,25 inspiring MIF-1 analogue synthesis to develop
therapeutics to treat Parkinson’s disease and depression. Aza-
pipecolyl MIF-1 analogue 40 was synthesized to study the
influence of conformational restriction of the prolyl residue on
biological activity by removal of the Boc group from 28b with
HCl gas in dichloromethane and aminolysis of ester 39 with
magnesium nitride in methanol (Scheme 5).26
The α-position of the cyclohexadiene ring in azapeptides 31a
and 32a was determined by hydrogenation with Raney nickel as
catalyst, which cleaved the N−N bond giving products with
molecular ion peaks of m/z [M + H]+ 456 and 380, respectively,
corresponding to fragments 36 and 37 (Scheme 4).
Scheme 4. Hydrogenolytic Cleavage and Mass Spectrometry
To Confirm Regioselectivity of Aza-allylglycines 31a and 32a
Pericyclic chemistry of azopeptides has provided effective
entry to azapeptides bearing aza-pipecolyl and aza-allylglycyl
residues without hydrazine chemistry using ionic intermediates.
Oxidation of carbamate protected aza-glycyl peptides followed by
Alder−ene products from reactions of azopeptide 13a with
cyclohexadiene and cycloheptatriene crystallized from EtOAc on
diffusion of hexane vapors. X-ray diffraction indicated α- and β-
substituted semicarbazides 30a and 35a, respectively (Figure 4).
In the solid state, α-cyclohexadienyl azapeptide 30a adopted
dihedral angle values indicative of a type I β-turn with
intramolecular 10-membered hydrogen bond between residues
i and i + 3 (Table 1). On the other hand, the torsion angles of β-
substituted azaglycine 35a were indicative of a type VIa-like β-
turn, albeit the urea ω-dihedral angle N-terminal to proline was
Scheme 5. Synthesis of Aza-pipecolylleucylglycinamide 40
C
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Letter
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ASSOCIATED CONTENT
* Supporting Information
The Supporting Information is available free of charge on the
ACS Publications website at DOI: 10.1021/acs.orglett.5b02723.
■
S
Experimental procedures, compound characterization
data, and NMR spectra for all new compounds (PDF)
X-ray data for 16a (CIF)
X-ray data for 28a (CIF)
X-ray data for 33a (CIF)
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AUTHOR INFORMATION
Corresponding Author
■
(14) Bourguet, C. B.; Sabatino, D.; Lubell, W. D. Biopolymers 2008, 90,
824.
*E-mail: lubell@chimie.umontreal.ca.
Notes
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The authors declare no competing financial interest.
ACKNOWLEDGMENTS
■
This research was supported by the Natural Sciences and
Engineering Research Council of Canada (NSERC). We thank
M. C. Tang, Sylvie Bilodeau, and Francine Belanger of the
Universite de Montreal centers for HR-MS analysis, NMR
́ ́
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