The synthesis and crystal structure of alpha-keto tetrazole-based dipeptide mimics

Article abstract of DOI:10.1016/S0040-4039(01)01101-7

Here we report the synthesis of a cis-dipeptide mimic N-Boc-Phe-[COCN₄]-Gly-OBn, 7, containing the non-hydrolysable α-keto tetrazole isostere and an unusual 2,5-disubstituted α-keto tetrazole-based peptidomimetic, 8. The incorporation of the no

Full text of DOI:10.1016/S0040-4039(01)01101-7

TETRAHEDRON
LETTERS
Pergamon
Tetrahedron Letters 42 (2001) 5641–5644
The synthesis and crystal structure of alpha-keto tetrazole-based
dipeptide mimics
Barnaby C. H. Maya,b,* and Andrew D. Abellb,*
aDepartment of Chemistry, University of Canterbury, Private Bag 4800, Christchurch, New Zealand
bDepartment of Cellular and Molecular Pharmacology, University of California San Francisco, San Francisco,
CA 94143-0450, USA
Received 15 March 2001; revised 18 June 2001; accepted 20 June 2001
Abstract—Here we report the synthesis of a cis-dipeptide mimic N-Boc-Phe-[COCN4]-Gly-OBn, 7, containing the non-
hydrolysable a-keto tetrazole isostere and an unusual 2,5-disubstituted a-keto tetrazole-based peptidomimetic, 8. The incorpora-
tion of the novel cis-amide bond isostere was achieved via direct alkylation of a precursor five-substituted (1H)-tetrazole. The
assignment of the resulting 1,5- and 2,5-tetrazoyl regiomers was based on the first reported X-ray structure analysis of an a-keto
tetrazole, compound 8. © 2001 Elsevier Science Ltd. All rights reserved.
We have embarked on a program to develop a range of
novel conformationally restricted amide bond isosteres
that incorporate a 1,5-disubstituted tetrazole.1 The tet-
razole ring has been shown to be an excellent mimic of
the cis-amide bond, and as such, this structural motif
can be used to pre-organise the amide bonds of pep-
tides, enzyme substrates and inhibitors into a cis-con-
formation.2 We have now extended this work with the
design and preparation of a novel conformationally
restricted a-keto tetrazole isostere [COCN4], 1, that
combines the conformational restriction of a 1,5-disub-
stituted tetrazole ring with the potency of a non-
hydrolysable a-keto amide isostere [COCONH], 2. The
a-keto amide functionality (e.g. 2) has found wide-
spread application as an isosteric replacement in
reversible inhibitors of the proteinase family of
enzymes. Amino acid derived a-keto amides have been
incorporated into inhibitors of a-chymotrypsin,3 cal-
pain,4 cathepsin B,5 pepsin6 and HIV protease.7 The
potency of the a-keto amide isostere has been
attributed to formation of a stabilised tetrahedral
adduct between the electrophilic carbonyl of the
isostere and catalytic residues of the proteolytic
enzyme.8 As an extension of this work, some simple
alpha-keto heterocycles have also been shown to inhibit
serine proteases.9 The a-keto tetrazole isostere, 1, is an
important addition to the amide bond analogues avail-
able to peptidomimetic chemists for incorporation into
enzyme inhibitors and conformational probes.
By far the most common route used to prepare a-keto
amide isosteres of type 2 is by oxidation of a precursor
a-hydroxy amino acid. In our earlier work,1 we synthe-
sised a tetrazole-based dipeptide mimic containing an
a-keto tetrazole isostere from a precursor dipeptide
containing an N-terminal a-acetoxy b-amino acid. The
amide linkage was then converted into the required
tetrazole on reaction with phosphorous pentachloride
and hydrazoic acid. The free hydroxyl group was then
revealed and finally oxidised to the a-keto substituent.
We now report a more expedient route to a-keto tetra-
zole-based dipeptide mimics.
It is known that five-substituted (1H)-tetrazoles can be
directly alkylated to yield 1,5-disubstituted and 2,5-dis-
ubstituted tetrazoles.10,11 However, difficulties are often
encountered when assigning the regiochemistry of the
resulting tetrazoyl products due to similar chemical
shifts between the 1,5- and 2,5-regioisomers. We postu-
lated that direct alkylation of a suitable (1H)-tetrazole
could be used for the preparation of the a-keto tetra-
zole isostere, 1. Using this approach we have success-
fully synthesised both the cis-dipeptide mimic
N-Boc-Phe-[COCN4]-Gly-OBn, (7), which contains an
a-keto tetrazole isostere and the unusual 2,5-disubsti-
* Corresponding authors.
0040-4039/01/$ - see front matter © 2001 Elsevier Science Ltd. All rights reserved.
PII: S0040-4039(01)01101-7

5642
B. C. H. May, A. D. Abell / Tetrahedron Letters 42 (2001) 5641–5644
disubstituted tetrazole would appear further upfield
than the Gly-a-H2 signal of the 2,5-disubstituted tetra-
zole. 13C NMR studies of 1- and 2-methyltetrazoles
have shown that the methyl and CN4 carbon atoms of
the 1,5-disubstituted tetrazole are more shielded than
the corresponding carbon atoms of the 2,5-disubsti-
tuted tetrazoles.18 Based on these reported observa-
tions, and the structural determination of 8 (see
discussion below), the tetrazole that was eluted first was
designated the 1,5-disubstituted tetrazole, 7, since it
showed a Gly-a-H2 1H resonance at l 5.47, and Gly-a-
CH2 and CN4 13C resonances at l 50.11 and l 148.09,
respectively. The more polar tetrazole, 8, had a Gly-a-
H2 1H resonance at l 5.53, and Gly-a-CH2 and CN4
13C resonances at l 53.76, and l 161.15, respectively.
More evidence for the stereochemical assignment of the
tetrazoyl regiomers, 7 and 8, was obtained by a het-
eronuclear multiple bond correlation (HMBC) experi-
ment. Compounds 7 and 8 show similar HMBC
correlations, however, only the 1,5-disubstituted tetra-
zole, 7, showed a correlation between the Gly-a-H2
protons and the CN4 carbon centre.
tuted tetrazoyl peptidomimetic, 8. Additionally, we
have been able to unequivocally assign the regioisomers
by NMR analysis and by solving a unique X-ray analy-
sis of the 2,5-disubstitiuted tetrazoyl product, 8.
The key steps in the synthesis of 7 and 8 involved
adding a C-terminal glycine residue by direct alkylation
of the tetrazole ring of 4 followed by oxidation of the
free hydroxyl group (Scheme 1). Tetrazole 4 was pre-
pared from the N-Boc-protected tetrazole, 3, itself pre-
pared in two steps according to the method of Satoh et
al.12 (49% overall yield) from commercially available
N-Boc-phenylalanine. A catalytic hydrogenation of the
N-Boc-protected tetrazole, 3, resulted in removal of the
benzyl methylether tetrazole protecting group, as
required, but also in reduction of the a-carbonyl group
(Scheme 1). Upon work-up, the reaction gave a mixture
of epimeric a-hydroxy (1H)-tetrazoles, 4a and 4b, in
90% yield, which was used without further purifica-
tion.13 Alkylation of the a-hydroxy (1H)-tetrazoles, 4a
and 4b, with benzyl bromoacetate in the presence of
N,N-diisoproplyethylamine (DIPEA) gave the epimeric
1,5-disubstituted tetrazoyl products, 5a and 5b, and
also the 2,5-disubstituted tetrazoyl products, 6a and 6b.
The products were purified, but not separated, by flash
column chromatography to give the regiomeric tetra-
zoles, 5 and 6, as a mixture in 60% yield.14 The a-
hydroxy tetrazoles, 5 and 6 were then oxidised, as a
mixture, with a solution of TEMPO,15 to give the
Single crystals of the 2,5-disubstituted tetrazole, 8, were
grown from methanol by slow evaporation. The struc-
ture of 8 was confirmed by an X-ray structure determi-
nation at 173(2) K and was satisfactorily refined (Fig.
1).19 To our knowledge this is the first structural deter-
1
regiomeric a-keto tetrazoles, 7 and 8 (3:4 by H NMR).
The tetrazoles, 7 and 8, were separated by flash column
chromatography, the less polar 1,5-disubstituted tetra-
zole, 7, eluting prior to the 2,5-disubstituted tetrazole,
8. The tetrazoles were obtained in 30 and 40% yields,
respectively.16
The structures of the regiomeric tetrazoles were
assigned on the basis of 1H and 13C NMR, and an
X-ray structure determination of 8. Based on previous
1H NMR studies of regiomeric tetrazoles,17 it would be
expected that the Gly-a-H2 1H resonance for the 1,5-
Figure 1. Crystal structure of the 2,5-disubstituted tetrazole,
N-Boc-Phe-[COCN4]-Gly-OBn, 8.
Scheme 1. (i) H2, 10% Pd/C, MeOH, rt, 24 h; (ii) DIPEA, BrCH2COOBn, CH2Cl2, rt, 24 h; (iii) TEMPO, KBr, NaOCl,
CH2Cl2/H2O, 0°C, 30 min.

B. C. H. May, A. D. Abell / Tetrahedron Letters 42 (2001) 5641–5644
5643
mination of an a-keto tetrazole-based peptidomimetic
and is the principle supporting data for the assignment
of the regiochemistry of the tetrazole products. In the
structure of 8 it is apparent that the tetrazole ring is
essentially planar with the ring torsion angles,
6. Hori, H.; Tasutake, A.; Minematsu, Y.; Powers, J. C. In
Peptides, Structure and Function; Deber, C. M.; Hruby,
V. J.; Kopple, K. D., Eds.; Pierce, Rockford, Illinois,
1985; p. 819.
7. Slee, D. H.; Laslo, K. L.; Elder, J. H.; Ollman, I. R.;
Gustchina, A.; Kervinen, J.; Zdanov, A.; Wlodawer, A.;
Wong, C.-H. J. Am. Chem. Soc. 1995, 117, 11867.
8. Angelastro, M. R.; Peet, N. P.; Bey, P. J. Org. Chem.
1989, 54, 3913.
C5ꢀN1ꢀN2ꢀN3,
N1ꢀN2ꢀN3ꢀN4,
N2ꢀN3ꢀN4ꢀC5,
N3ꢀN4ꢀC5ꢀN1, N4ꢀC5ꢀN1ꢀN2 being −0.3(4), 0.0(4),
0.3(4), −0.6(4) and 0.5(4)°, respectively. The tetrazole
ring of 8 showed a mean deviation from the plane of
,
0.002 A. The bond lengths of the tetrazole ring are all
9. Edwards, P. D.; Wolanin, D. J.; Andisik, D. W.; Davis,
similar, with the C5ꢀN1, N1ꢀN2, N2ꢀN3, N3ꢀN4,
M. W. J. Med. Chem. 1995, 38, 76.
10. Bavetsias, V.; Bisset, G. M. F.; Kimbell, R.; Boyle, F. T.;
Jackman, A. L. Tetrahedron 1997, 53, 13383.
N4ꢀC5, being, 1.345(4), 1.325(4), 1.351(3), 1.327(3),
,
and 1.352(4) A, respectively. The a-keto carbonyl bond
sits in the same plane as the tetrazole ring, with the
O18ꢀC17ꢀC5ꢀN4 and O18ꢀC17ꢀC5ꢀN4 torsion angles
being −178.1(3) and −0.1 (5)°, respectively. The core
isostere [C17···C6] is essentially planar, with a mean
11. Boyle, F. T.; Crook, J. W.; Matusiak, Z. S. UK Pat.
GB2272217A1, 1994; Chem. Abstr. 1995, 122, 160665.
12. Satoh, Y.; Moliterni, J. Synlett 1998, 528.
13. 4a and 4b: 1H NMR (300 MHz, CD3OD, for the mixture)
l 1.23, 1.26 (9H, s, Boc-(CH3)3), 2.90 (2H, AB system,
lA=2.76, lB=3.03, dd, J=6.9, 13.2 Hz, Phe-b-H2), 4.07,
4.12 (1H, m, Phe-a-H), 5.01 (1H, d, J=6.6 Hz, NH),
,
deviation from the plane of 0.013 A. The C17ꢀC5 and
N2ꢀC6 bonds are offset by −10°. The phenylalanine
ring is essentially planar and sits in a flagpole position
over the tetrazole heterocycle.
5.08, 5.13 (1H, bs, Phe-a-H), 7.17–7.28 (5H, m, ArH). 13
C
NMR (75 MHz, CD3OD, for the mixture) l 28.36, 28.82
(Boc-(CH3)3), 38.37, 39.52 (Phe-b-CH2), 57.77, 59.42
(Phe-a-CH), 67.22, 69.01 (Phe-CHOH), 80.50, 80.55
(Boc-C(CH3)3), 127.46, 127.67, 129.43, 129.63, 130.50
(ArCH), 139.38, 139.39 (ArC), 157.51 (CN4), 159.99,
160.02 (Boc-CO). ES MS 320.1721 C15H22N5O3 (MH+)
requires m/z 320.1723.
In this letter we have presented a short route to some
new conformationally restricted peptidomimetics, the
design of which is based on novel a-keto 1,5- and
2,5-disubstituted tetrazole [COCN4] amide bond
isosteres. We have also unequivocally assigned struc-
tures to the resulting 1,5- and 2,5-disubstituted tetrazole
isomers, a problem that is often encountered in tetra-
zole-based chemistry, using a combination of NMR
and X-ray crystallography.
14. 5a, 5b, 6a, and 6b: A solution of 1H-tetrazole as a
mixture of 4a and 4b (1.0 equiv., 110 mg, 0.35 mmol) in
dry CH2Cl2 (5 mL) was prepared in a flame-dried flask
under Ar. DIPEA (3.0 equiv., 130 mg, 1.04 mmol, 175
mL) was added dropwise to the stirred solution at rt.
After 5 min benzyl bromoacetate (2.0 equiv., 160 mg,
0.70 mmol, 110 mL) was added dropwise and the reaction
was stirred for 24 h. The reaction was diluted with ethyl
acetate (20 mL), washed with 10% aqueous HCl (2×10
mL), 1 M aqueous NaOH (2×10 mL), saturated aqueous
NaCl (10 mL), dried (MgSO4), filtered and evaporated.
The crude reaction product was purified by flash column
chromatography (40% ethyl acetate/petroleum ether) to
give a-hydroxy tetrazoles, 5a, 5b, 6a and 6b (127 mg,
Acknowledgements
The work was supported by a Royal Society of New
Zealand Marsden grant.
References
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77%), which were not separated. Selected H NMR (300
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(CH3)3), 3.04 (2H, m, Phe-b-H2), 4.20 (1H, m, Phe-a-H),
4.96 (1H, d, J=6.6 Hz, NH), 5.06 (1H, d, J=3.0 Hz,
Phe-CHOH), 5.14 (1H, d, J=5.1 Hz, Phe-CHOH), 5.20
(2H, bs, Bn-H2), 5.41 (2H, bs, Gly-a-H2), 7.17–7.36 (10H,
m, ArH). TLC (analytical, 40% ethyl acetate/petroleum
ether) Rf=0.17.
15. Harbeson, S. L.; Abelleira, S. M.; Akiyama, A.; Barrett,
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1
16. 7: (41 mg, 30%), eluted first as a clear oil. H NMR (300
MHz, CDCl3) l 1.35 (9H, s, Boc-(CH3)3), 3.22 (2H, AB
system, lA=3.07, lB=3.37, dd, J=8.1, 14.1 Hz, Phe-b-
H2), 5.12 (1H, d, J=6.6 Hz, NH), 5.21 (2H, d, J=4.8 Hz,
Bn-H2), 5.42 (1H, m, Phe-a-H), 5.47 (2H, d, J=3.0 Hz,
Gly-a-H2), 7.13–7.38 (10H, m, ArH). 13C NMR (75
MHz, CDCl3) l 27.97 (Boc-(CH3)3), 37.02 (Phe-b-CH2),
50.11 (Gly-a-CH2), 59.48 (Phe-a-CH) 68.14 (Bn-CH2),
80.25 (Boc-C(CH3)3), 124.03, 127.06, 128.38, 128.55,
128.68, 129.18 (ArCH), 134.14, 135.13 (ArC), 148.09
5. Peet, N. P.; Burkhart, J. P.; Angelastro, M. R.; Giroux,
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D. J. Med. Chem. 1990, 33, 394.

5644
B. C. H. May, A. D. Abell / Tetrahedron Letters 42 (2001) 5641–5644
(CN4), 154.00 (Boc-CO), 164.71 (Gly-CO), 189.05 (Phe-
19. Crystallographic data for 8: C24H27N5O5, M 465.51, mp
120–121°C, crystal dimensions 0.55×0.26×0.02 mm,
orthorhombic, a=6.140 (3), b=16.264 (7), c=23.523
CO). TLC (analytical, 30% ethyl acetate/petroleum
ether) Rf=0.39; IR (CDCl3, cm−1), 3392, 3324, 3215,
1756, 1632, 1603, 1528, 1497, 1230; ES MS 466.2097
C24H28N5O5 (MH+) requires m/z 466.2090. Further elu-
tion gave the 2,5-regioisomer, 8 (65 mg, 48%), as a
white solid. Mp 120–121°C; [h]2D0 −18.3 (c=0.00010,
methanol). 1H NMR (300 MHz, CDCl3) l 1.39 (9H, s,
Boc-(CH3)3), 3.19 (2H, AB system, lA=3.06, lB=3.33,
dd, J=6.9, 13.8 Hz, Phe-b-H2), 5.24 (1H, d, J=6.4 Hz,
NH), 5.29 (2H, s, Bn-H2), 5.53 (2H, s, Gly-a-H2), 5.58
(1H, m, Phe-a-H), 7.05–7.34 (10H, m, ArH). 13C NMR
(75 MHz, CDCl3) l 28.18 (Boc-(CH3)3), 37.93 (Phe-b-
CH2), 53.76 (Gly-a-CH2), 58.70 (Phe-a-CH) 58.57 (Bn-
CH2), 80.11 (Boc-C(CH3)3), 127.05, 128.52, 128.60,
128.74, 128.95, 129.34 (ArCH), 134.01, 135.24 (ArC),
155.00 (Boc-CO), 161.15 (CN4), 164.03 (Gly-CO),
189.32 (Phe-CO). TLC (analytical, 30% ethyl acetate/
petroleum ether) Rf=0.29; IR (CDCl3, cm−1), 3395,
3307, 3215, 1756, 1649, 1610, 1533, 1500; ES MS
466.2096 C24H28N5O5 (MH+) requires m/z 466.2090.
,
(10) A, h=90 (2), i=90 (2), k=90 (2)°, V=2349.0 (17)
3
,
A , space group P2(1)2(1)2(1), Z=4, F(000)=984,
D
calcd=1.316 mg/m3, absorption coefficient 0.096 mm−1
,
q range for data collection 2.14–26.45, index ranges
−75h57, −205k518, −295l529, data/restraints/
parameters 4837/0/307, goodness of fit on F2 was 0.794,
final R indices [I>2|(I)] R1=0.0448, wR2=0.0714, R
indices (all data) R1=0.1525, wR2=0.0904, largest dif-
ference peak and hole 0.174 and −0.355 e A−3. A
,
unique data set was measured at 173(2) K within
2qmax=57° limit (6 scans). Of the 30663 reflections
obtained, 4837 were unique (Rint=0.1593) and were
used in the full-matrix least-squares refinement. The
structure was solved by direct methods. Hydrogen
atoms were fixed in idealised positions. All non-hydro-
gen atoms were refined with anisotropic atomic dis-
placement parameters. Neutral scattering factors and
anomalous dispersion corrections for non-hydrogen
atoms were taken from Ibers and Hamilton. Crystallo-
graphic data for the structure in this paper have been
deposited with the Cambridge Crystallographic Data
Centre; deposition number CCDC 157463.
17. (a) Raap, R.; Howard, J. Can. J. Chem. 1969, 47, 813;
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.