Practical biocatalytic desymmetrization of meso-N-heterocyclic dicarboxamides and their application in the construction of aza-sugar containing nucleoside analogs

Article abstract of DOI:10.1039/c2cc18012j

Amidase-catalyzed desymmetrization of meso-N-heterocyclic dicarboxamides under very mild conditions provided a highly efficient and practical method for the preparation of enantiomerically pure carbamoyl-substituted heterocyclic amino acids that were unique and versatile platforms for the construction of both antipodes of aza-sugar containing nucleoside analogs.

Full text of DOI:10.1039/c2cc18012j

View Article Online / Journal Homepage / Table of Contents for this issue
Chemical Communications
www.rsc.org/chemcomm
Volume 48 | Number 29 | 11 April 2012 | Pages 3465–3564
ISSN 1359-7345
COMMUNICATION
Wang et al.
Practical biocatalytic desymmetrization of meso-N-heterocyclic dicarboxamides and their application in the
construction of aza-sugar containing nucleoside analogs

Dynamic ArticVleiewLiAnrktiscle Online
ChemComm
Cite this: Chem. Commun., 2012, 48, 3482–3484
www.rsc.org/chemcomm
COMMUNICATION
Practical biocatalytic desymmetrization of meso-N-heterocyclic
dicarboxamides and their application in the construction of aza-sugar
containing nucleoside analogsw
Peng Chen,a Ming Gao,a De-Xian Wang,a Liang Zhaob and Mei-Xiang Wang*b
Received 22nd December 2011, Accepted 28th January 2012
DOI: 10.1039/c2cc18012j
Amidase-catalyzed desymmetrization of meso-N-heterocyclic
dicarboxamides under very mild conditions provided a highly
efficient and practical method for the preparation of enantio-
merically pure carbamoyl-substituted heterocyclic amino acids
that were unique and versatile platforms for the construction of
both antipodes of aza-sugar containing nucleoside analogs.
for instance, has been shown to catalyze enantioselective
biotransformation of a large number of structurally varied
racemic amides to produce highly enantiopure carboxylic acid
and amide products.8,9 Surprisingly, biocatalytic enantio-
selective desymmetrization of meso-dicarboxamides remains
largely unexplored.10,11 On the basis of easy availability of
meso-dicarboxamide compounds, high catalytic efficiency and
enantioselectivity of amidases, we envisioned that the amidase-
catalyzed desymmetrization of meso-dicarboxamides would
provide a novel method for the synthesis of enantioenriched
amido-substituted carboxylic acid compounds, invaluable
chiral entities in organic synthesis.
Enantioselective desymmetrization reactions of meso compounds
constitute an elegant strategy for the synthesis of chiral products.
Being different from kinetic resolution, desymmetrization of meso
compounds in the presence of a chiral catalyst provides the enabling
technology to achieve a maximum yield of 100% with excellent
enantiocontrol. Furthermore, the method generally furnishes highly
enantiopure compounds that are not easily accessible by other
means. Ring opening reactions of meso epoxides,1 aziridines,2
carboxylic anhydrides3 and bicyclic hydrazines4 catalyzed by
either chiral Lewis acid catalysts or organocatalysts, for example,
have been shown to generate various functionalized organic
compounds of high enantiomeric purity. Owing to high catalytic
efficiency, selectivity and environmentally benign reaction condi-
tions, hydrolytic enzymes including lipases, esterases and acylases
have been extensively studied in desymmetric acylation of meso-
diols and hydrolysis of meso-diesters, leading to the formation of
diverse chiral building blocks that are essential for the synthesis
of natural products and bioactive compounds.5
Chiral N-heterocyclic compounds such as pyrrolidine and
piperidine derivatives occur widely as the core structures in natural
products and in synthetic pharmaceuticals.12 Functionalized
chiral pyrrolidine and piperidine compounds are also important
entities in asymmetric enamine catalysis.13 Apart from L-proline,
most of the enantiopure functionalized pyrrolidine and piperidine
derivatives are obtained from multi-step synthesis.12,13 We report
herein highly efficient and scalable synthesis of chiral pyrrolidine,
piperidine and dihydropyrrole compounds from enantio-
selective biocatalytic desymmetrization of meso-N-heterocyclic
dicarboxamides. To the best of our knowledge, it represents the
first example of amidase-catalyzed enantioselective desymmetri-
zation of meso-N-heterocyclic dicarboxamides. The resulting
biocatalytic products have also been demonstrated as valuable
platforms for the construction of both antipodes of aza-sugar
containing nucleoside analogs.
Amidases [3.5.1.4] are a type of hydrolytic enzymes catalyzing
the conversion of primary amides into carboxylic acids with the
release of ammonia. So far a large number of amidases and
microorganisms that contain amidases have been reported, and
some of them have been used extensively in the kinetic resolution
of racemic amides.6 Rhodococcus erythropolis AJ270,7 a nitrile
hydratase and amidase-containing microbial whole cell catalyst,
We initiated our study with the examination of biotransfor-
mation of meso-N-heterocyclic dicarboxamide 1. To facilitate
the isolation and the detection of products, the benzylation on
both secondary amine and carboxylic acid groups was executed
using benzyl bromide with the aid of K2CO3 (Scheme 1). Under
very mild conditions such as in neutral phosphate buffer at
30 1C, Rhodococcus erythropolis AJ270 was found able to
catalyze hydrolysis of meso-pyrroline-2,5-dicarboxamide 1a
efficiently. Complete conversion was observed within 1 h, and
product 2a was obtained in 90% yield. Most remarkably,
the amidase involved in Rhodococcus erythropolis AJ 270
exhibits excellent enantioselectivity against the substrate,
yielding enantiopure product 2a (499.5% ee).
a Beijing National Laboratory for Molecular Sciences, CAS Key
Laboratory of Molecular Recognition and Function, Institute of
Chemistry, Chinese Academy of Sciences, Beijing 100190, China
b The Key Laboratory for Organic Phosphorus Chemistry and
Chemical Biology (Ministry of Education), Department of
Chemistry, Tsinghua University, Beijing 100084, China.
E-mail: wangmx@mail.tsinghua.edu.cn; Fax: +8610 6279 6761;
Tel: +8610 6279 7527
w Electronic supplementary information (ESI) available: Experimental
details, 1H and 13C NMR spectra of products. CCDC 853593. For ESI
and crystallographic data in CIF or other electronic format see DOI:
10.1039/c2cc18012j
meso-2,5-Dihydro-1H-pyrrole-2,5-dicarboxamide 1b, the
compound bearing a carbon–carbon double bond, also acted
c
3482 Chem. Commun., 2012, 48, 3482–3484
This journal is The Royal Society of Chemistry 2012

View Article Online
Scheme 2 A large scale biocatalytic preparation of enantiomer 3.
Fig. 1 X-Ray molecular structure of 2aꢀHCl. A water molecule is
omitted for clarity.
at 30 1C in neutral phosphate buffer (250 mL) for 12 h led
straightforwardly to the formation of 19 g of pure enantiomer 3
(94% yield), which was isolated by precipitation and ion-exchange
column purification (Scheme 2) (see ESIw).
The structures of all products are supported by spectroscopic
data and microanalysis (see ESIw). The absolute configuration
of 2R,5S-5-carbamoylpyrrolidine-2-carboxylic acid product 2a
was determined by the single crystal X-ray diffraction analysis
of 2aꢀHCl saltz which was obtained conveniently from crystal-
lization by means of diffusion of diethyl ether into a mixture of
2a in aqueous HCl and ethanol (Fig. 1).
Scheme 1 Biocatalytic desymmetrization of 1a–d.
as an excellent substrate towards the amidase of Rhodococcus
erythropolis AJ270, and it underwent highly efficient biocatalytic
desymmetrization to give a single enantiomeric product 2b in
96% yield (Scheme 1). To our delight, the amidase involved in
Rhodococcus erythropolis AJ 270 exhibits excellent catalytic
efficiency and enantioselectivity against the six-membered
heterocyclic dicarboxamide substrate, meso-piperidine-2,6-
dicarboxamide 1c, producing enantiopure product 2c
(499.5% ee) in an almost quantitative yield. Only when the
substrate was expanded to the seven-membered meso-azepane-
2,7-dicarboxamide 1d, were a slightly low chemical yield
(74%) and diminished enantioselectivity (63.0% ee) obtained
(Scheme 1). High reaction velocity and excellent enantioselectivity
of biocatalytic desymmetrization of meso-dicarboxamide substrates
1a–c, which were similarly observed in the kinetic resolution of
racemic amino amides,14,15 may imply strong and highly enantio-
selective binding of the amidase towards secondary amino amide
functionality. Although the details of the mechanism of biocatalysis
at the molecular level await further investigation, lower catalytic
efficiency and enantiocontrol for 1d are most probably attributable
to its enlarged ring size and flexible conformational structure.
As a microbial whole cell biocatalyst, Rhodococcus erythropolis
AJ270 was found to be robust and tolerant to high concentration
of meso-dicarboxamide substrates and products, allowing
the scaled-up synthesis of chiral products. To demonstrate
the practical preparative biocatalysis, the amidase-catalyzed
desymmetrization of N-heterocyclic meso-dicarboxamides in the
multigram scale was conducted. Thus, incubation of 1a (20 g)
with Rhodococcus erythropolis AJ270 cells (2 g wet weight)
Functionalized enantiopure pyrrolidine, dihydropyrrole and
piperidine derivatives produced from biocatalytic desymmetri-
zation reactions are conceivably invaluable chiral building
blocks for the synthesis of natural products such as alkaloids
and their derivatives. In addition, pyrrolidine and particularly
dihydropyrrole assembled with cis-configured 2,5-functional
groups would render them as unique platforms for the construc-
tion of aza-sugar containing nucleoside analogs. Furthermore,
the facile chemical interconversions of the functional groups at
2- and 5-positions would provide accesses to both antipodes of
aza-sugar containing nucleoside analogs that are of significance
in the study of chemotherapeutic agents.16
To demonstrate the synthetic potential of biocatalytic
desymmetrization products, we implemented the practical
synthesis of the nucleoside-like compounds. As illustrated in
Scheme 3, Cbz-protection on nitrogen followed by esterifica-
tion of the carboxylic acid moiety of 3 gave product 4 in an
excellent yield. Dehydration of amide using SOCl2 in DMF
at 0 1C produced nitrile 5 in 90% yield. 1,3-Dipolar cyclo-
addition reaction between the cyano group and azide17 allowed
introduction of a tetrazole ring at the anomeric position.
Reduction of both carbamate and ester led to the formation
of aza-sugar containing nucleoside product 7 in high yield. To
synthesize the enantiomer of 7, compound 4 was readily
switched to its enantiomer ent-4 by means of functional group
transformations under practical conditions. For example,
treatment of ester 5 with LiOH at ambient temperature afforded
c
This journal is The Royal Society of Chemistry 2012
Chem. Commun., 2012, 48, 3482–3484 3483

View Article Online
Z = 1, m(Mo-Ka) = 0.71073; T = 173(2) K, full-matrix least-squares
refinement on F2 converged to structure Flack = ꢁ0.05 (6), RF = 0.0505
[I 4 2s(I)], 0.0522 (all data) and Rw(F2) = 0.1212 [I 4 2s(I)], 0.1232
(all data), goodness of fit 1.064. CCDC 853593.
1 For a review, see: C. Schneider, Synthesis, 2006, 3919.
2 For a review, see: M. Pineschi, Eur. J. Org. Chem., 2006,
4979.
3 For reviews, see: (a) S.-K. Tian, Y. Chen, J. Hang, L. Tang,
P. Mcdaid and L. Deng, Acc. Chem. Res., 2004, 37, 621;
(b) I. Atodiresei, I. Schiffers and C. Bolm, Chem. Rev., 2007,
107, 5683; (c) J. B. Johnson and T. Rovis, Acc. Chem. Res.,
2008, 41, 327.
4 For a review, see: C. Bournaud, F. Chung, A. P. Luna, M. Pasco,
G. Errasti, T. Lecourt and L. Micouin, Synthesis, 2009, 869.
5 For a leading review, see: (a) E. Garcia-Urdiales, I. Alfonso and
V. Gotor, Chem. Rev., 2005, 105, 313 and references cited therein.
For added examples, see: (b) A. Goswami and T. P. Kissick,
Org. Process Res. Dev., 2009, 13, 483; (c) U. Zutter, H. Iding,
P. Spurr and B. Wirz, J. Org. Chem., 2008, 73, 4895;
PLE-catalyzed hydrolysis of N-benzyl-pyrrolidine-2,5-dicarboxylic
diester has been reported to give half the ester product either with
moderate ee value or in low chemical yield. For examples, see:
(d) M.-a. Kurihara, K. Kamiyama, A. Kobayashi and S. Ohno,
Tetrahedron Lett., 1985, 26, 5831; (e) F. Bjorkling, J. Boutelje,
H. Hjalmarsson, K. Hult and T. Norin, J. Chem. Soc., Chem.
Commun., 1987, 1041.
6 For reviews on amidases in organic synthesis, see: (a) T. Sugai,
T. Yamazaki, M. Yokoyama and H. Ohta, Biosci., Biotechnol.,
Biochem., 1997, 61, 1419; (b) L. Martinkova and V. Kren, Biocatal.
Biotransform., 2002, 20, 73.
7 (a) A. J. Blakey, J. Colby, E. Williams and C. O’Reilly, FEMS
Microbiol. Lett., 1995, 129, 57; (b) R. O’Mahony, J. Doran,
L. Coffey, O. Cahill, G. W. Black and C. O’Reilly,
Antonie van Leeuwenhoek, 2005, 87, 221.
8 For reviews, see: (a) M.-X. Wang, Top. Catal., 2005, 35, 117;
(b) M.-X. Wang, Chimia, 2009, 63, 331; (c) M.-X. Wang,
Top. Organomet. Chem., 2011, 36, 105.
Scheme 3 Synthesis of aza-sugar containing nucleoside analogs 7 and
ent-7.
acid 8 which was easily converted into amide 9. The subsequent
Pinner reaction took place at ꢁ15 1C to yield desired ent-4.
Following the same procedures aforementioned for the prepara-
tion of 7, ent-7 was obtained from ent-4 (Scheme 3, see also ESIw).
In summary, we have shown for the first time the highly
efficient and scalable synthesis of enantiopure functionalized
pyrrolidine, dihydropyrrole and piperidine derivatives from the
amidase-catalyzed desymmetrization of meso-N-heterocyclic
dicarboxamides. The synthetic potential of the biotransformation
of chiral nitrogenous heterocycles has been demonstrated
preliminarily in the construction of both antipodes of aza-sugar
containing nucleoside analogs. The amidase-catalyzed desymmetri-
zation protocol would provide an enabling methodology for the
production of diverse enantiopure functional compounds that
are not easily available by other chemical means. Studies on
the understanding of the mechanism of amidase in enantio-
selective desymmetrization of dicarboxamides and its synthetic
applications are being actively pursued and the results will be
reported in due course.
9 For added examples, see: (a) D.-H. Leng, D.-X. Wang, J. Pan,
Z.-T. Huang and M.-X. Wang, J. Org. Chem., 2009, 74, 6077;
(b) D.-H. Leng, D.-X. Wang and M.-X. Wang, Org. Biomol. Chem.,
2010, 8, 4736.
10 Only biotransformation of meso-cyclohex-4-ene-1,2-dicarboxamide
was mentioned briefly in a communication. K. Matoishi, A. Sano,
N. Imai, T. Yamazaki, M. Yokoyama, T. Sugai and H. Ohta,
Tetrahedron: Asymmetry, 1998, 9, 1097.
11 For examples of enantioselective biotransformations of prochiral
diamides, see: (a) Z.-L. Wu and Z.-Y. Li, J. Org. Chem., 2003,
68, 2479; (b) M. Yokoyama, M. Kashiwagi, M. Iwasaki,
K.-i. Fuhshuku, H. Ohta and T. Sugai, Tetrahedron: Asymmetry,
2004, 15, 2817; (c) L.-B. Zhang, D.-X. Wang and M.-X. Wang,
Tetrahedron, 2011, 67, 5604.
12 Comprehensive Heterocyclic Chemistry III, ed. A. R. Katritzky,
C. A. Ramsden, E. F. V. Scriven and R. J. K. Taylor, Elsevier,
2008, ch. 3.03 in vol. 3 and ch. 7.05 in vol. 7.
13 For reviews, see: (a) S. Mukherjee, J. W. Yang, S. Hoffmann and
B. List, Chem. Rev., 2007, 107, 5471; (b) P. M. Pihko, I. Majander
and A. Erkkila, Top. Curr. Chem., 2010, 291, 29.
¨
14 (a) M.-X. Wang and S.-J. Lin, Tetrahedron Lett., 2001, 42, 6925;
(b) M.-X. Wang and S.-J. Lin, J. Org. Chem., 2002, 67, 6542;
(c) M.-X. Wang, S.-J. Lin, J. Liu and Q.-Y. Zheng, Adv. Synth. Catal.,
2004, 346, 439; (d) M.-X. Wang, J. Liu, D.-D. Wang and Q.-Y. Zheng,
Tetrahedron: Asymmetry, 2005, 16, 2409.
We thank Ministry of Science and Technology (Grant
No. 2009CB724704), National Natural Science Foundation of
China (Grant No. 20820102034), Chinese Academy of Sciences
and Tsinghua University for financial support.
15 D.-Y. Ma, D.-X. Wang, J. Pan, Z.-T. Huang and M.-X. Wang,
J. Org. Chem., 2008, 73, 4087.
16 (a) J. Benditt, Science, 1993, 260, 1253; (b) M. Ferrero and
V. Goto, Chem. Rev., 2000, 100, 4319; (c) E. Ichikawa and
K. Kato, Curr. Med. Chem., 2001, 8, 385; (d) D. Wang,
Y.-H. Li, Y.-P. Wang, R.-M. Gao, L.-H. Zhang and X.-S. Ye,
Bioorg. Med. Chem., 2010, 19, 41 and references cited therein.
17 A. Hartikka and P. I. Arvidsson, Eur. J. Org. Chem., 2005,
4287.
Notes and references
z Crystallographic data for 2aꢀHCl: C20H23ClN2O3.23, Mr = 378.53,
triclinic, P1, a = 5.614 (2) A, b = 8.720 (3) A, c = 10.053 (4) A,
a = 78.575 (11), b = 82.289 (12), g = 87.570 (15), V = 478.0 (3) A3,
c
3484 Chem. Commun., 2012, 48, 3482–3484
This journal is The Royal Society of Chemistry 2012