Expedient synthesis of N-Z-pyroglutamyl-amino acid derivatives

Article abstract of DOI:10.1016/j.bmcl.2007.07.052

N-Z-Pyroglutamyl pseudopeptides 3a-c are shown to be conveniently prepared from glutamyl-bis-Bt 1a by cyclization of an N-terminal glutamic acid residue. Structures are supported by 2D NMR studies and by comparison with the same products prepared by direc

Full text of DOI:10.1016/j.bmcl.2007.07.052

Bioorganic & Medicinal Chemistry Letters 17 (2007) 6000–6002
Expedient synthesis of N-Z-pyroglutamyl-amino acid derivatives
a,
a
a
b
*
Alan R. Katritzky, Parul Angrish, Ekaterina Todadze and Ion Ghiviriga
a
Center for Heterocyclic Compounds, Department of Chemistry, University of Florida, Gainesville, FL 32611-7200, USA
b
Department of Chemistry, University of Florida, USA
Received 10 April 2007; revised 16 July 2007; accepted 17 July 2007
Available online 19 August 2007
Abstract—N-Z-Pyroglutamyl pseudopeptides 3a–c are shown to be conveniently prepared from glutamyl-bis-Bt 1a by cyclization of
an N-terminal glutamic acid residue. Structures are supported by 2D NMR studies and by comparison with the same products pre-
pared by direct coupling of the C-terminus activated N-pGlu 1b and free amino acids 2a–c.
Ó 2007 Elsevier Ltd. All rights reserved.
Cellular mechanisms use regulation at different stages of
diverse functions. DNA to mRNA transcription con-
trol, mRNA to protein translation control, and further
protein post-translation modification are important
examples in the regulation of numerous cellular func-
tives are very late antigen-4 (VLA-4) antagonists de-
signed to inhibit the vascular cell adhesion molecule
(VCAM)/VLA-4 interaction and thus treat inflamma-
1
4,15
tory diseases such as asthma.
1
tions and activities. Post-translational modification of
Now, we report a convenient one-pot preparation of N-
Z-pyroglutamyl pseudopeptides derived from the cycli-
zation of the amino terminal glutamic acid residue
(Scheme 1, Route A). Thus, we show that peptide cou-
a protein is one of the later steps in protein biosynthesis
for many proteins. Post-translational modifications of
amino acids extend the range of functions of a protein
by attaching amino acids with other biochemically ac-
tive functional groups such as acetate, phosphate, lipids
or carbohydrates. Other post-translational changes af-
fect an amino acid more directly as in citrullination or
1
6
pling of N-Z-Glu-diBt 1a with diverse L-amino acids
2a–c (Ala, Phe and Val) in aqueous acetonitrile
(CH CN/H O) in the presence of Et N for 1 h (Scheme
3
2
3
1
6–18
1)
HCl gave 3a–c in 58–88% yields in high purity without
followed by washing the crude products with 4 N
2
–7
the formation of pyroglutamic acid (pGlu).
chromatography (Table 1).
The formation of pGlu occurs through cyclization of the
amino terminal residue (glu or gln) to pGlu inside the
cell prior to the activation of the completed protein.
In the present paper the structures of products 3a–c are
rigorously proved by 2D NMR (Fig. 1). Thus for 3a, the
cross-peaks in the gHMBC spectrum between the meth-
ylene protons at 2.43, 2.37, 2.26, 1.86 and the carbon at
174.3 indicate that the carbon at 174.3 is bound to that
at 31.5. Similar cross-peaks between the protons at 2.26,
1.86, 4.63 and the carbon at 171.4 indicate that this car-
bon is next to 59.4. The proton at 4.63 couples with the
carbon at 174.3 which confirms the 5-oxoproline moiety.
Other couplings confirm the presence of the second ami-
no acid moiety (alanine) and the coupling of protons at
8.51, 4.17 with carbon 171.4 indicates that this amino
acid is bound to proline. No coupling between 4.63
and 151.2 was seen, but the Z fragment must be con-
nected to the nitrogen in proline because these are the
only valences left unpaired.
1
In nature, simple and complex pseudopeptides partici-
pate in various biological processes. Much effort has
been focused on the synthesis of pseudopeptides as ago-
nists and antagonists in medical applications. The pres-
ence of pGlu is important for the biological activity of
1
,2,8–11
various proteins and peptides.
releasing hormone (TRH), L-pGlu-L-His-Pro-NH , is a
Thyrotropin-
2
tripeptide responsible for maintaining thyroid-stimulat-
1
ing hormone (TSH) levels in the anterior pituitary.
Pyroglutamic acid, the first residue of TRH, is responsi-
1
2,13
ble for at least half of the peptide’s binding energy.
Further, N-benzylpyroglutamyl-L-phenylalanine deriva-
Keywords: Protein; Protein biosynthesis; Functional groups; Acetate;
Phosphate; Lipids; Carbohydrates; Citrullination; Pyroglutamic.
1
6
*
Corresponding author. Tel.: +1 352 392 0554; fax: +1 352 392
199; e-mail: katritzky@chem.ufl.edu
In a previous paper from our group, the product from
the coupling reaction between Z-L-Glu-diBt 1a (1 equiv)
9
0
960-894X/$ - see front matter Ó 2007 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2007.07.052

A. R. Katritzky et al. / Bioorg. Med. Chem. Lett. 17 (2007) 6000–6002
6001
Scheme 1. Preparation of 3a–c using Route A.
Table 1. Preparation of 3a–c using Route A (Scheme 1) and Route B
(
Scheme 4)
a
Yield (%)
Entry Amino
acid
Product
Mp (°C)
Route A Route B
1
2
3
L-Ala 2a Z-pGlu-Ala 3a 58
L-Phe 2b Z-pGlu-Phe 3b 88
L-Val 2c Z-pGlu-Val 3c 71
55
81
70
205–207
173–174
154–156
Scheme 3. Preparation of 8.
a
Isolated yield.
We show that our methodology offers two fast and con-
venient routes (A, Scheme 1; B, Scheme 4) to prepare
pGlu pseudopeptides in good yields and high purity as
compared with literature methods. Preparations of
pseudopeptides of type 3 from the corresponding esters
or salts have been achieved using coupling reagents
and phenylalanine 2b (2 equiv) was reported as Z-L-Glu-
L-Phe-OH (see structure 5t in Ref. 16) We now know
that this was erroneous, because of a regrettable mistake
in the reading of 1D H NMR and C NMR. We also
assigned an incorrect structure to Z-L-Glu-L-di-Val-OH
1
13
1
9
20
including (i) DCC/HOBt/Et N, (ii) DEPC/Et N,
3 3
1
4,15
12
(
5u in Ref. 16) due to a similar error.
(iii) HBTU/DIEA,
(iv) EDC/HOBt/NEM. Other
methods are (v) active ester derived from 2,4,5-trichloro-
2
1
13
Recently, we reported the preparation of dipeptides
using C-a- and C-c-activated glutamic acid under varied
conditions leading to the formation of products 5 and 7
phenol or ethyl chloroformate, and (vi) a one-pot
Ugi-4-center-3-component reaction both in solution
phase and under microwave irradiation.
2
2
23
1
7,18
as shown in Scheme 2.
1
However, we now find that
when 6 is stirred in aqueous acetonitrile in the presence
8
In conclusion, we have demonstrated the preference in
glutamic acid derivatives of activated a-COOH over
activated c-COOH to undergo coupling thereby leading
to the formation of pyroglutamic dipeptides. These re-
sults were further supported by the preferred reactivity
route of activated c-COOH of glutamic acid in the
absence and presence of an amino acid with protected
a-COOH.
of Et N (but with no amino acid added), 6 cyclized to
3
form 8 (Scheme 3).
We also prepared 3a–c by direct coupling (Route B) of
Z-L-pGlu-Bt 1b with diverse amino acids 2a–c (Ala,
Phe, and Val) (Scheme 4). Routes A (Scheme 1) and B
(
Scheme 4) gave comparable yields of 3a–c (Table 1).
2
2
3
.43 2.26
.37 1.86
1.5 22.6
2.38
2.38
2.24
1.80
2.38 1.84
2.44 2.29
30.9 22.1
8
.51
8.58
8.37
31.7 22.7
4.66
1
74.5
173.4
172.6
4
5
.63
H
4.17
9.4 48.2
H
4.46
54.2
N
4.79
58.4
H
4.15
57.0
1
74.3
174.3
59.5
173.8
12.56
12.77
COOH 12.66
N
COOH
COOH
N
171.1
1
71.4
171.6
7.23
O
N
O
N
O
N
2.03
29.7
1
29.8
1
51.2
151.1
138.0
150.4
O3
3
2
7.3
.02
.89
7.26
128.9
O
O
17.7
1.21
O
O
O
O
O
O
17.8
0.84
19.0
0.82
5
5
.18
.12
5.1267.6
5.04
1
27.2
7.26
5.18
5.14
6
7.7
66.9
1
36.2
136.4
135.5
1
28.2
128.7
7.38 - 7.29
128.3
7.31
1
28.7
127.4
7.36
127.8
7.33
7
.35
7.33
1
29.1
129.1
.33
128.3
7.33
7
3a
3b
3c
1 13
Figure 1. H and C chemical shifts in compounds 3a–c, in DMSO-d
6
at 25 °C.
Scheme 2. Preparation of natural (5) and unnatural (7) dipeptides.

6002
A. R. Katritzky et al. / Bioorg. Med. Chem. Lett. 17 (2007) 6000–6002
Scheme 4. Preparation of 3a–c using Route B.
Supplementary data
10. Curran, T. P.; Marcaurelle, L. A.; O’Sullivan, K. M. Org.
Lett. 1999, 1, 1225.
1
1
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C.; Marshall, G. R.; Moeller, K. D. Bioorg. Med. Chem.
Synthetic details and compound characterization data
for 1b, 3a–c, and 8 is available free of charge via the
Internet at http://www.elsevier.com. Supplementary
data associated with this article can be found, in the on-
line version, at doi:10.1016/j.bmcl.2007.07.052.
2
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3. Sievertsson, H.; Chang, J. K.; Folkers, K. J. Med. Chem.
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