Nitrilases catalyze key step to conformationally constrained GABA analogous γ-amino acids in high optical purity

Article abstract of DOI:10.1021/jo070776j

(Chemical Equation Presented) Five- and six-membered carbocyclic γ-amino acids were prepared in high enantiomeric purity by nitrilase-mediated transformation of hitherto unreported γ-amino nitriles. The nitrilases investigated reveal a strong enantioprefe

Full text of DOI:10.1021/jo070776j

elaborate synthetic procedures for enantiomerically pure cyclic
GABA analogues.
Nitrilases Catalyze Key Step to Conformationally
Constrained GABA Analogous γ-Amino Acids in
High Optical Purity
Surprisingly, methods to prepare enantioenriched 3-ACPA
and 3-ACHA can scarcely be found in the literature. The
preparation of both enantiomers of cis-3-ACHA was achieved
via classical fractional crystallization of the diastereomeric
L-ornithine and brucine salts.7 Both isomers of 3-oxo-CPA were
obtained by resolution of their brucine adducts, following several
transformation steps to either enantiomer of cis- and trans-3-
ACPA.8 The cis enantiomers were also prepared by resolution
of their (-)-1-phenylethylammonium salts.9 cis-(-)-3-ACPA
was also isolated as a degradation product from the antiviral
antibiotic amidinomycin.10,11 In addition to these long-estab-
lished methods, approaches to cis-3-ACPA comprise multiple-
step chirospecific syntheses starting from aspartic acid,12
asymmetric allylic alkylation of 4-azido-1-benzoyloxycyclopent-
2-ene,13 and the asymmetric reductive amination of (()-3-
oxocyclopentanecarboxylic esters.14 In fact, few methods took
advantage of the benefits of biocatalysis. Thus, only the
enantiomers of cis-3-ACPA were obtained by esterase- and
lipase-catalyzed desymmetrization of meso cis-1,3-cyclopentane
dicarboxylic esters11,15 and by lactamase-catalyzed kinetic
resolution of a bicyclic lactam.16 However, the enantioselective
synthesis of trans-3-ACPA as well as either enantiomer of cis-
and trans-3-ACHA has not yet been accomplished using the
great resolution potential of biocatalysts.
For several years we have been interested in developing
efficient chemoenzymatic methods for the synthesis of non-
proteinogenic amino acids.17,18 In this context, we present here
our novel and practical access to enantioenriched γ-amino acids
from racemic amino nitriles (Figure 1) by the use of nitrilase.
Nitrilases are hydrolases (EC 3.5.5.1) capable of enantio-
selectively hydrolyzing a nitrile group via several steps, albeit
only the final product, the carboxylic acid, is released.19 In the
past, these enzymes have increasingly been applied for the
enantioselective preparation of acids20 because no intermediate
product of hydrolysis (amide) is released from the catalytic site,21
in contrast to the nitrile hydratase/amidase pathway. Incon-
Margit Winkler, Astrid C. Knall, Martin R. Kulterer, and
Norbert Klempier*
Institute of Organic Chemistry, Graz UniVersity of Technology
Stremayrgasse 16, A-8010 Graz, Austria
klempier@tugraz.at
ReceiVed April 13, 2007
Five- and six-membered carbocyclic γ-amino acids were
prepared in high enantiomeric purity by nitrilase-mediated
transformation of hitherto unreported γ-amino nitriles. The
nitrilases investigated reveal a strong enantiopreference for
cis-isomers (up to 99% ee), whereas trans-isomers were
available in up to 86% ee. The biocatalytic enantioselective
syntheses of cis-3-aminocyclohexanecarboxylic acid (3b),
trans-3-aminocyclohexanecarboxylic acids (4b, 6b, 8b) as
well as trans-3-aminocylopentanecarboxylic acid (2b) are
hereby reported for the first time.
γ-Amino butyric acid (GABA) is the major inhibitory
neurotransmitter in the mammalian central nervous system
(CNS).1 As a highly flexible molecule it can participate in many
low-energy conformation binding processes. In recent times,
the application of conformationally restricted GABA mimics
(mainly cyclic compounds containing a rigid backbone2) has
contributed to a better understanding in GABA neuroreceptor
research.3 For example, 3-aminocyclopentanecarboxylic acid (3-
ACPA) isomers were recognized to be particularly efficient
stereomeric probes for GABA binding site topography. 3-Amino-
cyclohexanecarboxylic acid (3-ACHA), however, was found to
act selectively as GABA uptake inhibitor.4 In addition, analogues
therefrom were investigated as therapeutic agents for a range
of CNS disorders.5,6 All these facts account for the need to
(7) Allan, R. D.; Johnston, G. A. R.; Twitchin, B. Aust. J. Chem. 1981,
34, 2231-2236.
(8) Allan, R. D.; Johnston, G. A. R.; Twitchin, B. Aust. J. Chem. 1979,
32, 2517-2521.
(9) (a) Milewska, M. J.; Polonski, T. Tetrahedron: Asymmetry 1994, 5(3),
359-362 (b) Amor´ın, M.; Castedo, L.; Granja, J. R. Chem. Eur. J. 2005,
11, 6543-6551.
(10) Nakamura, S.; Karasawa, K.; Yonehara, H.; Tanaka, N., Umezawa,
H. J. Antibiotics 1961, 14(Ser. A), 103-106.
(11) Cheˆnevert, R.; Lavoie, M.; Courchesne, G.; Martin, R. Chem. Lett.
1994, 93-96.
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58, 2369-2376.
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568-572.
(1) (a) Krogsgaard-Larsen, P. Med. Res. ReV. 1988, 8, 27-56 (b)
Andersen, K. E.; Sorensen, J. L.; Lau, J.; Lundt, B. F.; Petersen, H.;
Huusfeldt, P. O.; Suzdak, P. D.; Swedberg, M. D. J. Med. Chem. 2001, 44,
2152-2163 and references therein.
(14) Sung, S.-Y.; Frahm, A. W. Arch. Pharm. 1996, 329, 291-300.
(15) Cheˆnevert, R.; Martin, R. Tetrahedron: Asymmetry 1992, 3(2), 199-
200.
(2) Simonyi, M. Enantiomer 1996, 1, 403-414.
(16) Evans, C.; McCague, R.; Roberts, S. M.; Sutherland, A. G. J. Chem.
Soc., Perkin Trans. 1 1991, 656-657.
(3) Chebib, M.; Johnston, G. A. R. Clin. Exp. Pharmacol. Physiol. 1999,
26, 937-940.
(17) Preiml, M.; Hillmayer, K.; Klempier, N. Tetrahedron Lett. 2003,
44, 5057-5059.
(4) Krogsgaard-Larsen, P.; Froelund, B.; Frydenvang, K. Curr. Pharm.
Res. 2000, 6, 1193-1209.
(18) Winkler, M.; Mart´ınkova´, L.; Knall, A. C.; Krahulec, S.; Klempier,
N. Tetrahedron 2005, 61, 4249-4260.
(5) Allan, R. D.; Johnston, G. A. R. Med. Res. ReV. 1983, 3, 91-118.
(6) Chebib, M.; Duke, R. K.; Allan, R. D.; Johnston, G. A. R. Eur. J.
Pharmacol. 2001, 430, 185-192.
(19) (a) Brenner, C. Curr. Opin. Struct. Biol. 2002, 12, 775-782. (b)
Kobayashi, M.; Shimizu, S. FEMS Microbiol. Lett. 1994, 120, 217-224.
10.1021/jo070776j CCC: $37.00 © 2007 American Chemical Society
Published on Web 08/23/2007
J. Org. Chem. 2007, 72, 7423-7426
7423

diastereoselectivity (30:1) by reductive amination using ben-
zylamine as nitrogen donor and H2/Pd/C in MeOH as the
reduction system.
Initially, all γ-amino nitriles (()-1a-8a (Figure 1) were
subjected to nitrilase-mediated kinetic resolutions on a screening
level to give acids 1b-8b. To estimate the relative activity,
the reactions were stopped after 18 h (for conversions and ee’s
see Table 1 in Supporting Information). The screening results
suggest a preference of the nitrilases for the cis- versus the trans-
isomers with respect to conversion as well as enantioselectivity.
Evidently, the enzymes are sensitive toward changes in ring
size; thus, five-membered carbonitriles react superiorly through-
out as compared to six-membered substrates. trans-Carbonitriles
were accepted exclusively by NIT-104 and NIT-107, except
benzoate (()-8a, which was not hydrolyzed by any of the
nitrilases. The transformation of amino nitrile (()-5a was
accompanied by a loss of enantioselectivity compared to the
tosylate (()-3a. Replacement of the toluenesulfonyl group in
cis-amino nitrile (()-3a by benzoate (7a) decreases the conver-
sions significantly (<18%). For example, NIT-106 gave the acid
3b in 99% ee at 50% conversion, 5b in only 38% ee at 8%
conversion, whereas (()-7a was not accepted as substrate at
all. The reason for this obstacle of the benzoyl protection is
unclear yet. With regard to different N-protecting groups, a clear
trend for all the enzymes cannot be deduced. NIT-104 was more
successful for 7b, and NIT-105 for 5b in high optical yield (99%
ee and 94% ee, respectively), although both ee values were
achieved at a point of low substrate conversion (Table 1 in
Supporting Information). NIT-106 generally exhibited rever-
se enantioselectivity to all other applied nitrilases. Essentially,
nitrilases NIT-101-NIT-103 and NIT-105 turned out to be
unsuitable catalysts for the transformation of (()-1a-(()-8a.
On the basis of these screening results, optimized transforma-
tion protocols were established to synthesize particular acids
on a preparative scale with the appropriate nitrilase in high
optical purity (Table 1). The reactions were monitored by HPLC
and stopped at the time of the maximal expected enantiopurity
of the acids.29 The yields given in Table 1 were determined
after the isolation by extraction and chromatographic purifica-
tion. The optical purities of the remaining nitriles and acids were
determined by enantiomeric separation using HPLC; the result-
ing ee’s are depicted in Table 1 (see also Experimental Section).
In agreement with the screening, nitrilases NIT-106 and NIT-
107 are the most efficient catalysts with respect to the present
compounds. Thus, cis-3-ACPA (+)-1b was prepared with the
use of NIT-104, whereas (-)-1b was produced by NIT-106 in
an enantiocomplementary manner in almost enantiopure form
(97% ee) close to the theoretical yield of a kinetic resolution.
The respective trans-isomer 2b was obtained in only 55% ee
by the same enzyme. All other nitrilases examined could not
enhance this result. NIT-106 revealed similar outstanding
selectivities in the transformation of six-membered aminonitrile
cis-(()-3a to (-)-3b in almost optical purity (>99% ee) and
in 29% isolated yield at a comparable conversion with regard
to that of cis-(()-1a (Table 1). The enzyme’s prerequisite for
both high catalytic activity and enantioselectivity is best given
by a 1,3-diequatorial conformation of the substituents, as present
FIGURE 1. Novel (()-γ-amino nitriles (a) for enzymatic transforma-
tions to enantioenriched γ-amino carboxylic acids (b); (only one
enantiomer is depicted); Ts ) toluenesulfonyl; Cbz ) benzyloxycar-
bonyl; Bz ) benzoyl
veniently, a serious restriction for the organic chemist when
using these biocatalysts was the high efforts encountered so far,
when carrying out the reactions in whole cell transformations.
Hence, the recent availability of “ready for use” biocatalysts22
simplifies the reaction protocol considerably.
To provide the hitherto unreported amino nitriles23 (()-1a-
8a, we designed the straightforward procedures outlined in
Scheme 1, including relevant modifications of reported ap-
proaches. Michael addition of cyanide to R,â-unsaturated cyclic
ketones gave 3-oxo-cyclopentanecarbonitrile (()-9 and 3-oxo-
cyclohexanecarbonitrile (()-10. We made considerable im-
provements (see Supporting Information) compared with the
poor yields achieved by the literature procedures for both five-
and six-membered ketonitriles.24-26 Subsequent NaBH4 reduc-
tion of the ketones27 afforded epimeric hydroxy nitriles in a
cis:trans-isomer ratio of 88:12 for six-membered24 (()-16 and
(()-18 as well as 61:39 for five-membered rings (()-15 and
(()-17, determined by GC/MS analysis. Displacement of the
mesylates of (()-15-(()-18 by azide, reduction, and final
protection gave the desired amino nitriles in Scheme 1.
However, the minor formation of the trans-isomers (()-15 and
(()-16 by the ketonitrile reduction renders this access unfavor-
able for cis-aminonitriles. We tested several conditions for
reductive amination and found NaCNBH3 to reduce the inter-
mediate imines very efficiently to yield (()-11-(()-14.28 The
ratio of diastereomers, however, was close to equal. The
diastereomers are separable on silica gel after application of
the respective protecting groups. Alternatively, we found that
cis-amino nitriles (()-1a and (()-3a can be obtained in high
(20) (a) Mylerova´, V.; Mart´ınkova´, L. Curr. Org. Chem. 2003, 7, 1-17;
for some representative examples see: (b) Brady, D.; Beeton, A.; Zeevaart,
J.; Kgaje, C.; van Rantwijk, F.; Sheldon, R. Appl. Microbiol. Biotechnol.
2004, 64, 76-85. (c) DeSantis, G.; Wong, K.; Farwell, B.; Chatman, K.;
Zhu, Z.; Tomlinson, G.; Huang, H.; Tan, X.; Bibbs, L.; Chen, P.; Kretz,
K.; Burk, M. J. J. Am. Chem. Soc. 2003, 125, 11476-11477. (d) Layh, N.;
Stolz, A.; Fo¨rster, S.; Effenberger, F.; Knackmuss, H.-J. Arch. Microbiol.
1992, 158, 405-411. (e) Effenberger, F.; Bo¨hme, J. Bioorg. Med. Chem.
1994, 2(7), 715-721 (f) Kakeya, H.; Sakai, T.; Sano, A.; Yokoyama, M.;
Sugai, T.; Ohta, H. Chem. Lett. 1991, 1823-1824.
(21) For some important exceptions see: Winkler, M.; Glieder, A.;
Klempier, N. Chem. Commun. 2006, 12, 1298-1300 and references therein.
(22) Nitrilase NIT-101-NIT-108; BioCatalytics, Inc. Pasadena, CA.
(23) The preparation of cis-3-aminocyclopentane carbonitrile was de-
scribed in a very recent patent: WO 2006/040625 A1.
(24) Willaert, J. J.; Lemie`re, G. L.; Dommisse, R. A.; Lepoivre, J. A.;
Alderweireldt, F. C. Bull. Soc. Chim. Belg. 1984, 93, 139-149.
(25) Mertes, M. P.; Ramsey, A. A.; Hanna, P. E.; Miller, D. D. J. Med.
Chem. 1970, 13, 789-794.
(26) Agosta, W. C.; Smith, A. B., III. J. Am. Chem. Soc. 1971, 93, 5513-
5520.
(29) For a kinetic resolution rationale see: (a) Faber, K. Biotransfor-
mations in Organic Chemistry, 4th ed.; Springer-Verlag: Heidelberg, 2000.
(b) Poppe, L.; Nova´k, L. SelectiVe Biocatalysis; VCH: Weinheim, 1992.
(c) Chen, C.-S.; Fujimoto, Y.; Girdaukas, G.; Sih, C. J. J. Am. Chem. Soc.
1982, 104, 7294-7299.
(27) Benedetti, F.; Berti, F.; Garau, G.; Martinuzzi, I.; Norbedo, S. Eur.
J. Org. Chem. 2003, 1973-1982.
(28) Borch, R. F.; Bernstein, M. D.; Durst, H. D. J. Am. Chem. Soc.
1971, 93, 2897-2904.
7424 J. Org. Chem., Vol. 72, No. 19, 2007

SCHEME 1. Synthesis of (()-γ-Amino Nitriles (only one enantiomer is depicted)a
a Reagents and conditions: a) KCN, Et3N‚HCl, MeOH/H2O, 20 °C; b) I) ammonium formate, MS 4 Å, MeOH, rt; II) NaCNBH3; c) TsCl, Et3N, CH3CN,
rt;30 d) NaBH4, MeOH, rt; e) MsCl, Et3N, CH2Cl2, 0 °C;30 f) NaN3, DMF, 60 °C; g) 1 bar H2, Pd/C, MeOH, rt; h) BzCl, pyridine, CH3CN, rt;30 i) I) benzyl
chloroformate, K2CO3, Et2O/H2O, 0 °C; II) rt.30
TABLE 1. Nitrilase Biohydrolysis of Racemic N-Protected
γ-Amino Nitriles (a) to Enantioenriched γ-Amino Acids (b), Isolated
Yields
isomer of 3-aminocyclohexanecarboxylic acid was achieved by
NIT-107. The hereby presented approach represents the first
enantioselective synthesis of trans-configured five- and six-
membered ring carbocyclic γ-amino acids and their respective
six-membered cis-diastereomers.
substrate enzyme
time
(h)
conversion
(%)
ee
(%)
yield
(%)
substrate enzyme
(mM)
(g/L)
1a
2a
3a
4a
6a
NIT-106
NIT-106
NIT-106
NIT-107
NIT-107
0.56
0.34
0.42
0.67
0.22
0.50
0.50
0.50
1.00
0.50
1.75
30
9
256
18.5
47
36
42
46
41
97
55
>99
86
45
36
29
46
35
Experimental Section
74
The enzymes NIT-101-NIT-108 were purchased from a com-
mercial supplier.22 For preparative reactions, the enzymes used were
delivered with the specifications: NIT-104 (3.7 U/mg solid), NIT-
106 (85.0 U/mg solid), and NIT-107 (2.8 U/mg solid). For HPLC
analysis a LiChrospher 100 RP-18e column (5 µm) was used. Chiral
analysis was carried out with an Astec Chirobiotic R column, a
Chromtech Chiral AGP 100.4 column (5 µm), a Daicel Chiralpak
AD-H (5 µm), and a Chiralcel OD-H column (5 µm).
Screening. For screening experiments, 1.0 mg of enzyme
preparation was dissolved in 497.5 µL of phosphate buffer (50 mM,
pH 8.00, 1 mM EDTA), and 2.5 µL of the substrate was added as
solution in MeOH or DMSO (40 mM) to give a final concentration
of 0.2 mM. The reactions proceeded at 30 °C in a Thermomixer at
1100 rpm. After 18 h, 500 µL of acetone was added. The reaction
vessels were centrifuged at room temperature and 13000 rpm for 5
min to remove precipitated proteins. The supernatant was analyzed
by RP-18 HPLC using a gradient of 0.1% H3PO4 and acetonitrile.
For ee determination on carbohydrate-based chiral stationary phases,
the products were extracted using ethyl acetate. The solvent was
removed and the residue dissolved in MeOH prior to chiral HPLC
analysis.
in cis-(()-1a and cis-(()-3a. Notably, the remaining nitriles
were also recovered in up to 98% enantiomeric excess. Different
from that, the respective trans-isomers 4b and 6b were obtained
in good yields exclusively by NIT-107, although in diminished
enantiomeric excess (86% and 74%, respectively). In this case,
nitrilase NIT-106 completely failed to catalyze the transforma-
tions of trans-(()-4a and trans-(()-6a.31
The absolute configuration of cis-1b and cis-3b was assigned
to be (1R,3S) by comparison of the HPLC-elution order with
the only available reference acid cis-(1R,3S)-3-ACPA. By
serendipity, this configuration matches that of the natural
product, amidinomycin.10,11 For a detailed description on the
assignment of the other acids see the Supporting Information.
With these results it became clear that NIT-106 acted highly
R-selective in contrast to all other nitrilases applied in this study,
which were S-selective throughout.
In summary, the nitrilase NIT-106-mediated hydrolysis of cis/
trans-3-aminocyclopentane/hexane carbonitriles offers an ef-
ficient method for the enantioselective synthesis of (1R,3S)-
cis-3-aminocyclopentanecarboxylic acid and its six-membered
analogue (1R,3S)-cis-3-aminocyclohexanecarboxylic acid in high
optical purity. In contrast, the best optical yield for the trans-
Preparative Scale Biotransformations. The commercial enzyme
preparation was dissolved in phosphate buffer (50 mM, pH 8.00, 1
mM EDTA32) in a round-bottomed flask. The substrate was added
as solution in DMSO (max 5 vol % of cosolvent). The reaction
(31) Because no (published) crystal structure for any of the nitrilases is
available now, such striking differences in substrate reactivity can not be
rationalized. The same is true for the differing substrate specificity of NIT-
107 towards amino nitriles 4a and 6a, resulting in remarkably different
reaction times, depending on the protecting group
(30) For standard conditions, see: Greene, T. W.; Wuts, P. G. M.
ProtectiVe Groups in Organic Synthesis, 3rd ed.; J. Wiley and Sons: New
York, 1999.
J. Org. Chem, Vol. 72, No. 19, 2007 7425

was stirred with a magnetic bar, and the temperature was adjusted
to 29-31 °C by the use of an oil bath. The reaction was monitored
by HPLC (conversion and enantiomeric excess, as described for
the screening experiments). After completion, the products were
isolated by extraction with ethyl acetate and purified by silica gel
chromatography using ethyl acetate/cyclohexane mixtures with
small portions of acetic acid. In cases where the protein content
prevented the organic layer to clearly separate from the aqueous
layer, the protein was removed by precipitation using (NH4)2SO4
and filtration through a plug of Celite.
Yield 35 mg of (+)-2b (36% at 36% conversion, ee ) 55%); [R]20
D
+0.2 (c 1.34, CH2Cl2) from 90 mg of (()-2a; 1H NMR (CDCl3) δ
1.44-1.51 (m, 1H), 1.73-1.81 (m, 2H), 1.88-1.95 (m, 1H), 1.97-
2.10 (m, 2H), 2.43 (s, 3H), 2.90 (m, 1H, H-1), 3.71 (m, 1H, J )
6.2 Hz, H-2), 5.07 (s, br, 1H, NH), 7.31 (d, 2H, J ) 7.6 Hz), 7.75
(2H, d, J ) 7.6 Hz), 8.45 (s, br, 1H, COOH); 13C NMR (CDCl3)
δ 21.8, 27.7, 33.5, 36.6, 41.4, 54.8, 127.3, 130.0, 137.5, 143.8,
181.3; IR (CaF) 3270, 1705, 1599, 1321, 1157 cm-1; Chiral
separation on Chirobiotic R, polar organic mode (MeOH/Et3N/
AcOH, 100:0.1:0.4), 0.80 mL/min, 30 °C.
cis-3-(Toluene-4-sulfonylamino)cyclohexanecarboxylic acid
(3b): white solid, mp 134-138 °C; NIT-106: Yield 55 mg of (-)-
cis-3-(Toluene-4-sulfonylamino)cyclopentanecarboxylic acid
(1b):33 white solid, mp 127-129 °C; NIT-104: Yield 65 mg of
(+)-(1S,3R)-1b (83% at 89% conversion, ee ) 17%), [R]20D +0.6
(c 1.09, CH2Cl2) from 73 mg of (()-1a; NIT-106: Yield 69 mg of
(-)-(1R,3S)-1b (45% at 47% conversion, ee ) 97%), [R]20D -24.6
(c 1.00, CH2Cl2) from 147 mg of (()-1a; 1H NMR (CDCl3) δ 1.59-
1.68 (m, 1H), 1.71 (tt, 1H, J ) 5.1, 1.7 Hz), 1.74-1.82 (m, 1H),
1.85-1.97 (m, 2H), 1.98-2.04 (m, 1H), 2.43 (s, 3H), 2.82 (m,
1H), 3.76 (m, 1H), 5.54 (s, br, 1H, NH), 6.41 (s, br, 1H, COOH),
7.29 (d, 2H, J ) 8.1 Hz), 7.75 (d, 2H, J ) 8.1 Hz); 13C NMR
(CDCl3) δ 21.8, 28.3, 33.7, 36.4, 41.6, 54.9, 127.3, 130.0, 138.0,
143.7, 181.7; m/z (EI) 201 (10), 173 (8), 155 (19), 91 (100), 67
(91); IR (CaF) 3238, 1705, 1598, 1322, 1158 cm-1; Chiral
separation on Chirobiotic R, polar organic mode (MeOH/Et3N/
AcOH, 100:0.1:0.4), 0.80 mL/min, 30 °C.
(1R,3S)-3b (48% at 50% conversion, ee ) 88%), [R]20 -45.0 (c
D
1.09, CH2Cl2) from 113 mg (()-3a; NIT-106: Yield 8.5 mg of
(-)-(1R,3S)-3b (29% at 42% conversion, ee >99%), [R]20D -43.6
(c 0.355, CH2Cl2) from 23 mg of (()-3a; 1H NMR (CDCl3) δ 1.07-
1.14 (m, 1H), 1.22-1.29 (m, 2H), 1.39-1.42 (m, 1H), 1.78-1.84
(m, 2H), 1.91 (m, 1H), 2.10 (m, 1H), 2.30 (m, 1H), 2.43 (s, 3H),
3.10-3.13 (m, 1H), 4.76 (s, br, 1H, NH), 6.24 (s, br, 1H COOH),
7.30 (d, 2H, J ) 8.1 Hz), 7.75 (d, 2H, J ) 8.1 Hz); 13C NMR
(CDCl3) δ 21.5, 23.9, 27.7, 33.4, 35.7, 41.9, 51.9, 126.9, 129.7,
138.1, 143.4, 179.4; m/z (EI) 215 (3), 186 (3), 173 (3), 155 (7),
124 (5), 91 (100); IR (CaF) 3271, 1705, 1599, 1321, 1157 cm-1
;
Chiral separation on AGP, 10 mM phosphate buffer pH 7.01, 0.80
mL/min, 15 °C.
Acknowledgment. We cordially acknowledge the NMR
operators Carina Illaszewicz and Hansjo¨rg Weber for their
assistance.
Supporting Information Available: Details of the preparation
of γ-amino nitriles, characterization data (1a-8a, 4b-6b, 1c-6c,
and 8c), enantiomer separation, determination of absolute configu-
rations, screening results (1a-8a), results of additional preparative
biotransformations (1b-4b and 6b) as well as copies of all relevant
NMR spectra. This material is available free of charge via the
Internet at http://pubs.acs.org.
trans-3-(Toluene-4-sulfonylamino)cyclopentanecarboxylic acid
(2b):33 white solid, mp 139-141 °C; NIT-106: Yield 72 mg (81%
at 90% conversion, ee ) 6%) from 83 mg of (()-2a; NIT-106:
(32) Addition of DTT was omitted for reasons outlined in ref 21
(33) The racemic compound was mentioned in: Berger, H.; Paul, H.;
Hilgetag, G. Chem. Ber. 1968, 101, 1525-1531. No spectral data are
included.
JO070776J
7426 J. Org. Chem., Vol. 72, No. 19, 2007