7748 J. Am. Chem. Soc., Vol. 122, No. 32, 2000
Moon et al.
[MB]
R
PCDMeO was collected by filtration and washed with 30 mL of
antibody with the highest peptidase activity reported to date.
Considering the difference in pH and temperature, the catalytic
activity of the PCD-based catalysts prepared in the present study
appears to be comparable to or better than that of the catalytic
antibody.
methanol and 30 mL of DMF. EPMA analysis indicated that the amount
of unsubstituted chloromethyl group is negligible.
MeO
â r
[(tren) ] PCD . The MB moiety attach to the PCD backbone was
MeO
disassembled by shaking [MB]
R
PCD
with 10 mL of concentrated
HCl mixed with 10 mL of acetone at 25 °C for 2 days and by washing
with 30 mL of acetone and 30 mL of methylene chloride. The color of
the resin turned to yellow from green as MB moieties were disas-
sembled. The amount of Cu(II) ion liberated during disassembly of
MB with acid was quantified by ICP-AES. From the amount of Cu(II)
ion, the content (R) of MB in [MB] PCD
Another feature of natural peptidases reproduced by the PCD
derivatives of the present study is the substrate selectivity.
Carboxypeptidase A, for example, is an exopeptidase recogniz-
ing terminal carboxylate ions of proteins. Chymotrypsin, trypsin,
and papain are some of endopeptidases that recognize side
chains with unique structural features in the vicinity of the
MeO
R
was calculated.
MeO
MeO
[(Cu(II)tren) ] PCD . To [(tren) ] PCD
â
r
â R
(1 g) stirred in a
1
scissile amide bonds. In this regard, the PCD derivatives with
methylene chloride solution (10 mL) of triethylamine (2.9 g, 29 mmol)
at room temperature for 3 h, 12 mL of DMF solution of CuCl (3.3
mM) was added and the mixture was stirred at room temperature
overnight. The resin of [(Cu(II)tren)
filtration and washed thoroughly with water and an aqueous buffer
active sites comprising three proximal Cu(II)tren units manifest
substrate selectivity toward amides containing proximal car-
boxylate ion.
2
MeO
â
]
R
PCD
was collected by
Binuclear or trinuclear metal complexes can be prepared
solution of pH 6 (0.01 M 4-morpholineethanesulfonic acid (mes)). After
42-45
through stepwise synthetic pathways.
For example, a
MeO
[(Cu(II)tren)
]
â R
PCD
(0.040 g) was shaken in a mixture of 1 mL of
trinuclear complex has been synthesized using calixarene as
concentrated HCl and 1 mL of acetone for 1 day at room temperature
at the speed of 165 rpm, the resin was washed with 10 mL of water
and the Cu(II) ion liberated was quantified by ICP-AES. From the
amount of Cu(II) ion thus obtained and the value of R estimated as
described above, the value of â was calculated.
N,N′-Phthaloyl-N′′-tert-butyloxycarbonyl-tren (Phth
To a solution of tren (1.0 g, 6.8 mmol) and Na CO (2.2 g, 20 mmol)
in 100 mL of water and 20 mL of DMF, a DMF solution (5 mL) of
N-carbethoxy phthalimide (3.0 g, 14 mmol) was added. After the
solution was stirred at room temperature for 3 h, di-t-butyl carbonate
44
scaffold. It would be very difficult, however, to prepare a three-
dimensional molecular device with several convergent catalytic
elements and to tune the geometry of the device by stepwise
synthesis to obtain a highly effective artificial enzyme. In this
regard, the novel strategy reported in the present study, transfer
of metal-chelating sites confined in a prebuilt cage to a
polymeric backbone, would be valuable in construction of active
sites of artificial multinuclear metalloenzymes.
2
BOC-tren).
2
3
(
2.2 g, 10 mmol) was added to the mixture and the resulting solution
was stirred for 2 h. Ethyl acetate (200 mL) was added to the mixture,
and the resulting solution was washed with 0.1 M NaHCO solution
twice and with saturated NaCl solution. After drying with MgSO , the
solvent was removed in vacuo. The product was recrystallized from
Experimental Section
Synthesis of Catalysts: PCD. This polymer was obtained by
suspension copolymerization of chloromethylstyrene (7:3 mixture of
m and p isomers) and divinylbenzene with benzoyl peroxide in the
presence of cyclohexane and poly(vinyl alcohol) in water as described
3
4
1
ethyl acetate-hexane, mp 149.5-150 °C. H NMR (300 MHz, CDCl
δ 1.35 (s, 9H), 2.84 (t, 2H), 2.88 (t, 4H), 3.12 (t, 2H), 3.72 (t, 4H),
: C, 63.44;
3
)
1
3
previously. The content of divinylbenzene was 2 mol % relative to
chloromethylstyrene.
5.00 (s, 1H), 7.27-7.77 (m, 8H). Anal. Calcd for C27
30 4 6
H N O
H, 6.04; N 10.77. Found: C, 64.02; H, 5.97; N, 11.06.
MB. Macrocyclic complex 1(ClO
4
)
4
2
‚2H O was synthesized as
2
4
MeO
reported previously.
MB] PCD. Attachment of MB to PCD to obtain [MB]
1 2
[(Cu(II)tren) ]1.7PCD . To a solution of Phth BOC-tren (0.067
[
r
R
PCD was
g, 0.13 mmol) in 20 mL of methylene chloride, trifluoroacetic acid
(0.15 g; 1.3 mmol) was added, and the resulting mixture was stirred at
room temperature for 30 min to remove the BOC group. The excess
trifluoroacetic acid was removed by evaporation in vacuo. The resulting
residue was dissolved in 50 mL of methylene chloride, to which PCD
(1 g) and N,N-diisopropylethylamine (0.40 mL; 2.3 mmol) were added.
The mixture was shaken at 25 °C for 5 days to attach the tren derivative
to PCD. After the resin was washed with 100 mL of water and 100
mL of acetone, it was suspended in a 1:1 (v/v) mixture (200 mL) of
DMF and methanol. After sodium methoxide (2.0 g; 37 mmol) was
added to the mixture, the mixture was shaken at 50 °C for 2 days to
replace chloro groups with methoxide. After the resin was washed with
50 mL of methanol and 50 mL of acetone, the mixture of the resin and
methylamine (2.0 mL; 3.0 mmol) in 50 mL of methanol was shaken at
50 °C for 3 days to remove the phthalimide-protecting group, and the
1
product ([(tren) ]1.7PCD ) was washed with 50 mL of methanol and
100 mL of acetone. Upon treatment of [(tren)
solution (6 mL) of 0.01 M CuCl , [(Cu(II)tren)
Removal of the phthalimide-protecting group was confirmed by Cu-
(II) binding to the resulting tren unit. The amount of tren moieties
carried out under nitrogen atmosphere with solvents degassed prior to
use in synthesis. PCD (0.5 g; 3.3 residue mmol) and NaH (0.050 g;
2
.1 mmol) were added to DMF (25 mL) taken from a fresh bottle and
the resin was swollen for 1 day at room temperature. NaH (0.020 g,
0
×
.83 mmol) was added to a DMF (5 mL) solution of MB (0.045 g; 4.0
10 mol) at room temperature and the mixture was stirred for 30
-
5
min and then the undissolved NaH powder was removed by filtration.
Upon treatment with NaH, the color of the solution turned to brown
from green. The brown solution of MB was added to PCD suspended
in DMF, and the resulting mixture was shaken for 2 days at 50 °C.
The speed of shaking employed in the synthesis of various derivatives
of PCD in the present study was 60 rpm unless noted otherwise. When
MB was attached to PCD, the resin became green. The green resin of
[MB]
R
PCD was collected by filtration and washed with 30 mL of
MeO
methanol, 50 mL of water, and 50 mL of acetone. When it was
attempted to vary the degree of cross-linkage of MB by PCD,
p-nitrobenzoic acid (50, 100, or 200 mol % relative to MB: 0.13 g
MeO
1
]
1.7PCD
with DMF
MeO
2
1
]
1.7PCD
was formed.
n-
MB was treated with 0.030 g NaH for generation of MB when 100
or 200 mol % p-nitrobenzoic acid was used) was added to the solution
n-
n-
of MB for partial protonation of the MB
.
attached to the resin was quantified by measuring the amount of Cu-
(II) ion released after treatment of [(Cu(II)tren) ]1.7PCD
MeO
MeO
with 6 N
[
MB]
with methoxide ion to obtain [MB]
prepared with 1.0 g PCD) with sodium methoxide (0.86 g, 16 mmol)
in 100 mL of 1:1 (v/v) DMF-methanol at 50 °C for 30 h. The resin of
r
PCD . The chloro groups of [MB]
R
PCD were substituted
1
MeO
MeO
HCl. EPMA analysis of [(Cu(II)tren)1]1.7PCD
indicated the amount
R
PCD
by shaking [MB] PCD
R
of unsubstituted chloromethyl group is negligible.
Synthesis of Substrates: N-Methyl Cinnamoyl Amide (3). This
compound was prepared from cinnamoyl chloride and methylamine
and was recrystallized from ethyl acetate-hexane, mp 111.5-112.5
°
(
(
42) Wall, M.; Hynes, R. C.; Chin, J. Angew. Chem., Int. Ed. Engl. 1993,
2, 1633.
43) Molenveld, P.; Engbersen, J. F. J.; Kooijman, H.; Speck, A. L.;
Reinhoudt, D. N. J. Am. Chem. Soc. 1998, 120, 6726.
44) Molenveld, P.; Engbersen, J. F. J.; Reinhoudt, D. N. Angew. Chem.,
Int. Ed. 1999, 38, 3189.
45) He, C.; Lippard, S. J. J. Am. Chem. Soc. 2000, 122, 184.
3
46
C (lit. 110-111 °C).
(
N-Cinnamoyl Glycine (4), N-Cinnamoyl â-alanine (5), and
(
N-Cinnamoyl γ-butyric Acid (6). To a mixture of the corresponding
(
(46) Briggs, L. B.; de Ath, G. E.; Ellis, S. R. J. Chem. Soc. 1942, 61.