Parallel Solid-Phase Synthesis and Evaluation of Inhibitors of Streptomyces coelicolor Type II Dehydroquinase

Article abstract of DOI:10.1021/jm030987q

A series of 1-substituted and 4-substituted benzyl analogues of the known inhibitor (1S,3R,4R)-1,3,4-trihydroxy-5-cyclohexene-1-carboxylic acid has been synthesized and tested as inhibitors of Streptomyces coelicolor type II dehydroquinase. The solid-phas

Full text of DOI:10.1021/jm030987q

J . Med. Chem. 2003, 46, 5735-5744
5735
P a r a llel Solid -P h a se Syn th esis a n d Eva lu a tion of In h ibitor s of Str eptom yces
coelicolor Typ e II Deh yd r oqu in a se
Concepcio´n Gonza´lez-Bello,*,† Emilio Lence,† Miguel D. Toscano,‡ Luis Castedo,† J ohn R. Coggins,§ and
Chris Abell‡
Departamento de Qu´ımica Orga´nica y Unidad Asociada al C.S.I.C., Facultad de Qu´ımica,
Universidad de Santiago de Compostela, 15782 Santiago de Compostela, Spain, University Chemical Laboratory,
Lensfield Road, Cambridge CB2 1EW, UK, and Biochemistry and Molecular Biology, University of Glasgow,
Glasgow G12 8QQ, UK
Received August 5, 2003
A series of 1-substituted and 4-substituted benzyl analogues of the known inhibitor (1S,3R,4R)-
1,3,4-trihydroxy-5-cyclohexene-1-carboxylic acid has been synthesized and tested as inhibitors
of Streptomyces coelicolor type II dehydroquinase. The solid-phase syntheses of 18 new
analogues are reported. The most potent inhibitor, 2-nitrobenzyloxy analogue 5i, has Ki of 8
µM, more than 30 times lower than the KM of the substrate and approximately 4 times more
potent than the original inhibitor. The binding modes of the synthesized analogues in the active
site were studied by molecular docking with GOLD 2.0.
In tr od u ction
dehydroquinase with 3 bound in the active site has
recently been solved.10 This complex identifies a number
of key interactions involved in inhibitor binding and
sheds light on aspects of the catalytic mechanism of the
enzyme. Also present in this structure was a molecule
of glycerol, originated from the enzyme storage buffer,
bound 3.7 Å away from the inhibitor 3. This fact was
used to design bifuntional inhibitors that straddle the
two binding sites identified in the crystal structure of
the enzyme.11 The important observation that com-
pounds with a double bond between C5-C6 bind in the
manner predicted for transition state mimics encour-
aged us to design the next generation of inhibitors.
In this paper we describe attempts to make more
potent inhibitors by incorporating binding interactions
onto the core structure 3. We describe the synthesis of
18 new analogues, 4a -i and 5a -i (Scheme 2), using
solid-phase organic synthesis (SPOS). The inhibition
studies and the molecular docking with these com-
pounds against S. coelicolor type II dehydroquinase are
also described.
The shikimate pathway is the biosynthetic pathway
to the aromatic amino acids phenylalanine, L-tryp-
tophan, and L-tyrosine, as well as precursors to the
folate coenzymes, alkaloids, and vitamins.1 The pathway
is present in bacteria, fungi, and plants and has been
recently discovered in apicomplexan parasites.2 Its
absence from mammals has made this pathway an
attractive target for the development of herbicides and
antimicrobial agents. The successful herbicide Glypho-
sate acts by specifically inhibiting the sixth enzyme on
the pathway.3
Dehydroquinase (3-dehydroquinate dehydratase) is
the third enzyme on the shikimate pathway and is
responsible for catalyzing the conversion of 3-dehydro-
quinic acid (1) to 3-dehydroshikimic acid (2) (Scheme
1). There are two forms of the enzyme, type I (e.g. from
Escherichia coli)4 and type II (e.g. from Streptomyces
coelicolor).5 The type I enzyme mechanism involves
covalent imine intermediates between the enzyme and
the substrate and proceeds with syn stereochemistry.6
In contrast, the type II reaction proceeds through an
enolate intermediate with overall anti stereochemistry.7
These mechanistic and stereochemical distinctions have
allowed us to design and synthesize compounds that are
specific to either the type I8 or type II9 enzymes.
Syn th esis of C-1 Su bstitu ted An a logu es. The
strategy used for making the analogues 4a -i involved
the initial preparation of the resin 8 (Scheme 3). This
was made from hydroxycarbolactone 7, which was
synthesized from benzoate 6 using our previously
reported protocol.12 Treatment of bromo-Wang resin13
with the sodium alkoxide of lactone 7 afforded the
lactone resin 8. Although the reaction can be carried
out using THF or DMF and at room temperature or 60
°C, the best results were obtained in DMF at 60 °C. The
gel-phase FT-IR spectrum of the ether lactone resin 8
The enolate intermediate in the type II reaction is
flattened relative to the 3-dehydroquinic acid (1). In
addition, a negative charge is localized toward the
enolate oxygen. Both of these features have been
exploited in the design of first-generation inhibitors.9
We have previously reported that analogue 3 (Scheme
2) showed a Ki of 30 µM for S. coelicolor type II
dehydroquinase. The crystal structure of S. coelicolor
showed the lactone stretching band at 1797 cm-1
.
Further evidence for the formation of 8 was obtained
from examination of the gel-phase 13C NMR spectrum.
Distinctive signals for the TBS group were observed at
25.6 and -3.1 ppm. In addition, the signal for the
methylene benzyl group moved from 34.0 ppm in bromo-
Wang resin to 71.2 ppm in the lactone resin 8.14
Deprotection of the tertiary hydroxyl group in 8 was
* Corresponding author. Tel: +34 981 563100. FAX: +34 981
595012. E-mail: cgb1@lugo.usc.es.
† Universidad de Santiago de Compostela.
‡ University Chemical Laboratory.
§ University of Glasgow.
10.1021/jm030987q CCC: $25.00 © 2003 American Chemical Society
Published on Web 11/22/2003

5736 J ournal of Medicinal Chemistry, 2003, Vol. 46, No. 26
Gonza´lez-Bello et al.
Sch em e 1. Conversion of 3-Dehydroquinic Acid (1) to 3-Dehydroshikimic Acid (2) Catalyzed by Type II
Dehydroquinase
Sch em e 2. S. coelicolor Type II Dehydroquinase
Sch em e 3. Parallel Solid-Phase Synthesis of C-1
Inhibitors
Analogues
achieved by treatment of the lactone resin 8 with TBAF
in THF at room temperature for 2 h. The gel-phase FT-
IR spectrum of hydroxylactone resin 9 showed a strong
band centered at 3414 cm-1. The complete deprotection
of the hydroxyl group was confirmed in the gel-phase
13C NMR spectra of 9.
Treatment of hydroxylactone resin 9 with sodium
hydride in DMF at 0 °C followed by treatment with an
excess of the corresponding benzyl bromide and heating
between 40 and 80 °C gave the corresponding ethers
10a -i.15 This reaction was highly dependent on the
nature of the benzyl bromide. Electron-donating groups
reacted faster and required lower reaction temperatures
(40 °C). Electron-withdrawing groups slowed the reac-
tion and required higher temperatures (80 °C). The
synthesis of the nitro derivatives was particularly
problematical, and the reaction proceeded very slowly.
The extent of conversion of each benzylation reaction
was monitored by gel-phase FT-IR spectroscopy follow-
ing the disappearance of the band at 3414 cm-1 corre-
sponding to the tertiary hydroxyl group of hydroxy-
lactone 9. The ether formation went to completion in
all cases, except for the nitrobenzyl analogues. Treat-
ment of the resins 10a -g with 50% TFA/DCM at room
temperature for 1 h afforded the corresponding lactones.
Basic hydrolysis and washing the aqueous layer with
diethyl ether to remove apolar impurities, followed by
treatment with Amberlite IR-120 (H+) ion-exchange
resin and lyophilization, gave the desired acids 4a -g
in good yield and excellent purities.

Inhibitors of S. coelicolor Type II Dehydroquinase
J ournal of Medicinal Chemistry, 2003, Vol. 46, No. 26 5737
Sch em e 4a
a
Reagents and Conditions: (i) TBAF, THF, 0 °C (92%); (ii) (1) HNa, DMF, 0 °C; (2) nitrobenzyl bromide, 15-crown-5, 80 °C [12 (41%);
13 (64%)]; (iii) KCN, MeOH, rt [15 (60%) and 14 (traces); 16 (23%) and 17 (63%)]; (iv) (1) LiOH, THF, rt, (2) Amberlite IR-120 (H+) [4h
(96%), 4i (78%)].
Sch em e 5. Parallel Solid-Phase Synthesis of C-4 Analogues
Because of the slow reaction of 9 with nitrobenzyl
bromide, the analogues 4h and 4i were eventually
obtained as minor components in mixtures with un-
alkylated 3. Consequently, we decided to prepare 4h and
4i by solution-phase synthesis. The four-step synthesis
is outlined in Scheme 4. Treatment of the silyl ether
derivative 6 with TBAF gave the desired tertiary alcohol
11 in excellent yield. Alkylation of the corresponding
alkoxide of 11 with 4-nitrobenzyl bromide afforded the
corresponding ether 12 in 41% yield (with 30% of
recovered starting material). The best yields for the
alkylation reaction were obtained when the reaction was
performed in DMF at 80 °C for 48 h in the presence of
a catalytic amount of 15-crown-5 ether. No significant
difference was observed if sodium or potassium hydride
was used. The benzoyl group was removed by treatment
of the lactone 12 with potassium cyanide in methanol
at room temperature to form the methyl ester 15 in 60%
yield and traces of lactone 14. Finally, conversion of
methyl ester 15 to the desired acid 4h was achieved by
ester hydrolysis, followed by treatment with Amberlite
IR-120 (H+) ion-exchange resin and lyophilization. A
similar strategy was used for the preparation of acid 4i
using 2-nitrobenzyl bromide.
Syn th esis of th e 4-Su bstitu ted An a logu es. The
4-substituted benzyl analogues 5a -g were prepared on
solid-phase as outlined in Scheme 5. The strategy used
for making analogues 5a -g required the initial prepa-
ration of the resins 19a ,b. Alkylation of bromo-Wang
resin with the corresponding sodium alkoxide of hy-
droxycarbolactone 11 in DMF at 80 °C afforded the
desired benzyl ether resin 18. The gel-phase FT-IR
spectrum of ether resin 18 showed two new stretching
bands centered at 1794 and 1718 cm-1 corresponding

5738 J ournal of Medicinal Chemistry, 2003, Vol. 46, No. 26
Gonza´lez-Bello et al.
Sch em e 6a
Ta ble 1. Inhibition Results for Assays with S. coelicolor Type
II Dehydroquinasea
compd
R
Ki (µM)
compd
R
Ki (µM)
4a
4b
4c
4d
4e
4f
4g
4h
4i
H
380 ( 30
375 ( 30
425 ( 40
260 ( 20
170 ( 15
5a
5b
5c
5d
5e
5f
5g
5h
5i
H
200 ( 20
70 ( 5
140 ( 10
130 ( 10
100 ( 10
4-F
3-F
2-F
4-F
3-F
2-F
4-CO2H
4-CH2OH 1330 ( 100
4-CN
4-NO2
2-NO2
4-CO2H
4-CH2OH >2000
4-CN
4-NO2
2-NO2
430 ( 40
980 ( 90
45 ( 5
330 ( 30
200 ( 20
8 ( 2
a
KM value of 250 µM was obtained at the assay conditions.
ing tertiary alcohol 23. Finally, treatment of lactone 23
with lithium hydroxide, followed by treatment with
Amberlite IR-120 (H+) ion-exchange resin and lyo-
philization, afforded the desired acid 5h . Acid 5i was
prepared by a similar reaction sequence using 2-nitro-
benzyl bromide.
Assa y Resu lts. The 18 acids 4a -i and 5a -i were
assayed in the presence of 3-dehydroquinic acid (1) for
their inhibitory properties against S. coelicolor type II
dehydroquinase. The inhibition data are summarized
in Table 1. A UV spectrophotometric assay was used to
measure the initial rate of product formation, detecting
the enone-carboxylate chromophore at 234 nm in
3-dehydroshikimic acid (2). The assays with 4e, 5e, 4g,
and 5g were performed at 260 nm due to the strong UV
absortion of these analogues at 234 nm. The Ki values
were obtained from Dixon plots (1/v vs [I]).
All the compounds were shown to be reversible
competitors of S. coelicolor type II dehydroquinase. The
4-substituted series was found to be generally more
potent than the 1-substituted series. However, there
was a general correlation across the two series. The
three isomeric fluorobenzyl derivatives showed potency
similar to or somewhat lower than the parent compound
(4a and 5a ) in both series. The 2-nitro derivatives were
the most potent compounds in each series. The 4-sub-
stituted analogue 5i had a Ki of 8 µM compared with
30 µM for the original inhibitor 3, and considerably
below the KM of 250 µM. The 4-carboxy benzyl deriva-
tives 4e and 5e were both potent, the 4-cyano analogues
4g and 5g less so. The 4-hydroxymethyl compounds 4f
and 5f were surprisingly poor inhibitors. The correlation
between the two series broke down for the 4-nitrobenzyl
derivatives, where the 1-substituted compound 4h was
considerably less potent than the 4-substituted analogue
5h .
Dock in g Stu d ies. To gain some insight into how the
inhibitors were binding to the active site of S. coelicolor
type II dehydroquinase, docking studies were carried
out using GOLD 2.0.16 The starting point for this study
was the crystal structure of the enzyme with the
anhydro inhibitor 3 bound in the active site.10 This
structure has the carboxyl group of 3 binding to the
backbone amides of Ile107 and Ser108 and the C-1
hydroxyl binding to His106. There was an additional
binding pocket close by that was occupied by a glycerol
molecule (from the buffer). The molecule of 3, the
glycerol molecule, and all the water molecules except
one (which is conserved in a series enzyme inhibitor
complexes)10 were removed from the structure. No
energy minimization was performed on the enzyme. The
structures of the inhibitors were prepared and energy-
a
Reagents and Conditions: (i) (1) HNa, DMF, 0 °C; (2) nitro-
benzyl bromide, Bu4NI, 15-crown-5, 80 °C [21 (24%); 22 (54%)];
(ii) TBAF, THF, 0 °C [23 (43%); 24 (71%)]; (iii) (1) LiOH, THF, rt,
(2) Amberlite IR-120 (H) [5h (74%), 5i (94%)].
to the new carbonyl groups. Examination of the gel-
phase 13C NMR spectrum showed distinctive signals at
73.6, 69.9, and 67.5 ppm.14 Deprotection of the benzoyl
group in resin 18 was achieved by treatment of 18 with
potassium cyanide in methanol at room temperature for
2 h. This gave the resin 19 in two forms: as the
carbolactone 19a (minor) and as the methyl ester 19b
(major). The gel-phase FT-IR spectrum of hydroxy resins
19a ,b confirmed the disappearance of the benzoyl ester
band at 1718 cm-1. The spectra showed a new strong
band at 1736 cm-1 corresponding to the methyl ester
and also the stretching band corresponding for the
lactone at 1792 cm-1. The alkylation of the resin 19a ,b
was achieved by heated the corresponding sodium
alkoxide of 19a ,b with the various benzyl bromides in
DMF in the presence of a catalytic amount of 15-crown-5
ether. The extent of conversion was followed by moni-
toring the disappearance of the band at 3596 cm-1
corresponding to the hydroxyl group in 19a ,b in the gel-
phase FT-IR spectra. In all the cases the ether formation
went to completion. Treatment of the resins 20a -g with
50% TFA/DCM at room temperature for 1 h afforded
the corresponding lactones. Finally, basic hydrolysis and
washing the aqueous layer with diethyl ether, followed
by treatment with Amberlite IR-120 (H+) ion-exchange
resin and lyophilization, afforded the desired acids 5a -g
in good yields and excellent purities.
It again proved problematical to make the nitrobenzyl
analogues on solid-phase, so they were synthesied in
solution as outlined in Scheme 6. Treatment of the
corresponding sodium alkoxide of 7 with 4-nitrobenzyl
bromide in DMF at 80 °C for 48 h in the presence of a
catalytic amount of 15-crown-5 ether afforded the cor-
responding ether 21 in 24% yield, with recovery of 40%
unreacted starting material. The silyl group was re-
moved by treatment with TBAF to yield the correspond-

Inhibitors of S. coelicolor Type II Dehydroquinase
J ournal of Medicinal Chemistry, 2003, Vol. 46, No. 26 5739
F igu r e 1. Comparison between position of 3 (purple) in the crystal structure of S. coelicolor type II dehydroquinase and the
docking results of the highest score solution of (a) ligand 5a (green), (b) ligand 4e (blue), and (c) ligand 5i (yellow).
minimized either using SYBYL6.5 and the Tripos force
field or using Gaussian 98W17 and AM1. All the inhibi-
tors were docked as their carboxylate anions and 25
independent GOLD runs were performed for each
ligand. The ChemScore scoring function was used.
The 4-substituted series 5a -i (with the exception of
5i) docked so that the anhydro ring of the inhibitor
occupied approximately the same site as was occupied
by 3 in the crystal structure of the enzyme inhibitor
complex,10 and the benzyl group occupied the pocket
that had previously been occupied by the glycerol
molecule (Figure 1a). The carboxyl and C-1 hydroxyl
groups of the inhibitors occupied the same binding
pocket as for 3, although the orientation of the actual
ring was twisted relative to the binding of 3. The key
binding interactions for the benzyl substituent appeared
to be π stacking against Tyr28 and electrostatic or
hydrogen-bonding interactions with Arg23. The potency
of the 4-carboxy and 4-nitro analogues 5e and 5h may
be due to this latter interaction. The poor binding of the
hydroxymethyl analogue 5f may be because of limited
space at the top of the binding pocket.
The 1-substituted series 4a -i bound in a generally
similar orientation to the 4-substituted series. The
anhydro ring of the inhibitor occupied approximately
the same site as was occupied by 3 in the crystal
structure of the enzyme-inhibitor complex, and the
benzyl group occupied the pocket that had previously
been occupied by the glycerol molecule (Figure 1b).
However, the different position of the benzyl substituent
on the anhydro ring led to some differences in the
binding relative to the 4-substituted series. These are
most clearly seen in a different orientation of the
anhydro ring, caused by the need to put the benzyl
group into the glycerol pocket. The carboxyl group still
bound into the carboxylate binding pocket, but in some
analogues the anhydro ring was conformationally flipped
so that the C-5 hydroxyl group can bind to His106. The
benzyl substitutent occupied a similar position to that
found for the 4-substituted series, with the same
interactions with Tyr28 and Arg23. The latter interac-
tion may again be responsible for the relative potency
of the 4-carboxybenzyl analogue 4e.
carboxylate binding site (Figure 1c). The anhydro ring
docked into the glycerol pocket, with the carboxylate
binding to Arg23. This result is surprising and may be
an artifact of the way the active site is prepared for the
docking (if more of the original water molecules are
retained, docking similar to that for the other 4-sub-
stitued analogues was observed).
These docking studies have proved useful in giving
some clues as to the possible binding orientations of the
analogues and have highlighted some possible interac-
tions that can be targeted in the next generation of
inhibitors. However, they need to be supported with
structural studies of enzyme-inhibitor complexes, and
these are underway.
Con clu sion s
Solid-phase organic synthesis and solution-phase syn-
thesis have been used to make a series of 1-benzylated
and 4-benzylated analogues of the known inhibitor 3.
The 4-substituted 2-nitrobenzyl analogue 5i was found
to have a Ki of 8 µM, making it one of the most potent
known inhibitor against any type II dehydroquinase.
Docking studies have identified two binding pockets
likely to be occupied by the inhibitor and provide some
rationale for the relative potencies of the inhibitors.
Exp er im en ta l Section
Gen er a l P r oced u r es. All starting materials and reagents
were commercially available and used without further puri-
fication. FT-IR spectra were recorded as NaCl plates or KBr
disks. [R]D values are given in 10-1 deg cm2 g-1 1H NMR
.
spectra (250, 300, and 500 MHz) and 13C NMR spectra (63,
75, and 100 MHz) were measured in deuterated solvents. J
values are given in hertz. NMR assignments were made by a
combination of 1 D, COSY, and DEPT-135 experiments. All
procedures involving the use of ion-exchange resins were
carried out at room temperature and used Mili-Q deionized
water. Amberlite IR-120 (H+) (cation exchanger) was washed
alternately with water, 10% NaOH, water, 10% HCl, and
finally water before use. Purity of the carboxylic acids was
analyzed by HPLC and NMR. HPLC was performed on a
preparative (300 mm × 16 mm) Bio-Rad Aminex Ion exclusion
HPX-87H organic acids column. The eluant used for these
columns was 75 mM aqueous formic acid, at a flow rate of 0.6
mL min-1
.
The one anomalous result from the docking study was
the binding predicted for the most potent inhibitor 5i,
the 4-substituted 2-nitrobenzyl analogue. This sug-
gested that the relative positions of the two rings was
reversed, so that the benzyl group occupied the binding
site occupied by 3, with the nitro group binding into the
Deh yd r oqu in a se Assa y. S. coelicolor type II dehydro-
quinase was purified as described previously,5 as a concen-
trated solution (1.5 mg mL-1) in potassium phosphate buffer
(50 mM, pH 7.0); DTT (1 mM) was filter-sterilized through a
0.2 µm filter and stored at 4 °C under which conditions it was
stable for at least 9 months. When required for assays, aliquots

5740 J ournal of Medicinal Chemistry, 2003, Vol. 46, No. 26
Gonza´lez-Bello et al.
25
1
of the enzyme stocks were diluted into water and buffer and
stored on ice.
[R]D +15° (c 0.7 in H2O); H NMR (250 MHz, D2O) δ (ppm)
7.31 (m, 5H), 5.93 (d, 1H, J 10.1), 5.82 (dd, 1H, J 10.1 and
1.8), 4.41 (s, 2H), 4.00 (dd, 1H, J 8.3 and 1.8), 3.77 (td, 1H, J
11.9, 8.3, and 3.5), 2.14 (dd, 1H, J 13.6 and 3.5), and 1.87 (dd,
1H, J 13.6 and 11.9); 13C NMR (100 MHz, D2O) δ (ppm) 178.1,
137.9, 134.2, 128.8, 128.8, 128.4, 127.3, 80.8, 72.6, 69.5, 67.7,
and 37.9; υmax (KBr)/cm-1 3434, 1714, and 1578; MS (CI) m/z
(%) 247 (MH+ - H2O); HRMS calcd for C14H15O4 (MH+ - H2O)
247.0970, found 247.0965.
Dehydroquinase was assayed in the forward direction by
monitoring the increase in absorbance at 234 nm in the UV
spectrum due to the absorbance of the enone-carboxylate
chromophore of 3-dehydroshikimic acid (2) (ꢀ/M-1 cm-1 12 000).
Standard assay conditions for type II dehydroquinase were pH
7.0 at 25 °C in Tris/HCl (50 mM). Each assay was initiated by
addition of the enzyme. Solutions of 3-dehydroquinic acid (1)
were calibrated by equilibration with type II dehydroquinase
and measurement of the change in the UV absorbance at 234
nm due to the formation of the enone-carboxylate chromo-
phore of 3-dehydroshikimic acid (2). In the case of acids 4e,
5e, 4g, and 5g, assays were performed at 260 nm.
Dock in g. The receptor and ligands were used as MOL2
files. Each ligand was docked using GOLD 2.0 in 25 inde-
pendent genetic algorithm (GA) runs, and for each of these a
maximum number of 100 000 GA operations was performed
on a single population of 50 individuals. Operator weights for
crossover, mutation, and migration in the entry box were used
as default parameters (95, 95, and 10, respectively), as well
as the hydrogen bonding (4.0 Å) and van der Waals (2.5 Å)
parameters. The position of the active site was introduced and
the radius was set to 15 Å, with the automatic active-site
detection on. The “flip ring corners” flag was switched on, while
all the other flags were off.
P r ep a r a tion of Resin 8. To a stirred solution of the alcohol
712 (1.4 g, 5.18 mmol) in dry DMF (20 mL) at 0 °C under inert
atmosphere was added sodium hydride (222 mg, 5.54 mmol,
ca. 60% in mineral oil). After 30 min at this temperature the
resultant suspension was added via cannula to a preswollen
PS-brominated Wang resin13 (1 g, ∼1.6 mmol/g, ∼1.6 mmol)
in dry DMF (17 mL) at 0 °C and under inert atmosphere. Then,
15-crown-5 ether (30 µL, 0.26 mmol) was added and the
resultant suspension was shook at 0 °C for 30 min and at room
temperature for 24 h. The resin was thoroughly washed with
DMF, DMF-water (3:1), THF, and dry DCM and dried under
high vacuum to afford pale yellow beads of 8 (1.02 g): gel-
phase FTIR (cm-1) 1797 and 1611; gel-phase 13C NMR (63
MHz, CDCl3) δ (ppm) 128.5, 118.7, 114.7, 73.7, 71.2, 69.9, 64.8,
40.4, 37.7, 25.6, and -3.1.
P r ep a r a tion of Resin 9. To preswollen resin 8 (1 g, ∼1
mmol) in dry THF (16 mL) at 0 °C and under inert atmosphere
was added a solution of tetrabuthylammonium fluoride in THF
(1.4 mL, ca. 1 M in THF). The resultant suspension was shook
for 2 h at room temperature. The resin was thoroughly washed
with THF, THF-5% HCl (3:1), THF, and dry DCM to afford
after drying in vacuo pale yellow beads of resin 9 (∼0.9 g):
gel-phase FTIR (cm-1) 3414, 1789, and 1609; gel-phase 13C
NMR (63 MHz, CDCl3) δ (ppm) 136.4, 129.5, 121.5, 115.3, 74.6,
71.1, 53.5, and 36.9.
(1R,3R,4R)-1-(4′-F lu or oben zyloxy)-3,4-d ih yd r oxycyclo-
h ex-5-en -1-ca r boxylic a cid (4b): 67% overall; retention time
26 min; [R]D +9° (c 1.0 in H2O); 19F NMR (282 MHz, D2O) δ
25
(ppm) -112.6 (tt, 1F, J 9.1 and 5.2); 1H NMR (250 MHz, D2O)
δ (ppm) 7.32 (dd, 2H, J 7.7 and 5.7), 7.04 (t, 2H, J 8.7), 5.91
(m, 2H), 4.40 (s, 2H), 4.03 (dd, 1H, J 8.2 and 1.2), 3.78 (ddd,
1H, J 11.6, 8.2, and 3.5), 2.15 (dd, 1H, J 13.6 and 3.5), and
1.90 (dd, 1H, J 13.6 and 11.6); 13C NMR (63 MHz, D2O) δ (ppm)
177.8, 162.7 (J 242), 134.8, 133.8, 131.1 (J 8), 127.0, 115.6 (J
21), 80.3, 72.7, 69.6, 67.2, and 38.0; υmax (KBr)/cm-1 3420, 1716,
and 1605; MS (CI) m/z (%) 265 (MH+ - H2O); HRMS calcd for
C
14H14O4F (MH+ - H2O) 265.0876, found 263.0880.
(1R,3R,4R)-1-(3′-F lu or oben zyloxy)-3,4-d ih yd r oxycyclo-
h ex-5-en -1-ca r boxylic a cid (4c): 63% overall; retention time
29 min; [R]D +11° (c 0.6 in H2O); 19F NMR (282 MHz, D2O)
25
δ (ppm) -111.8 (dt, 1F, J 9.6 and 5.6); 1H NMR (250 MHz,
D2O) δ (ppm) 7.61 (m, 1H), 7.43-7.26 (m, 3H), 6.23 (d, 1H, J
10.1), 6.14 (dd, 1H, J 10.1 and 1.8), 4.72 (s, 2H), 4.31 (dt, 1H,
J 8.2 and 1.8), 4.08 (ddd, 1H, J 12.2, 8.2, and 3.7), 2.44 (ddd,
1H, J 13.6, 3.0, and 1.1), and 2.19 (dd, 1H, J 13.6 and 12.2);
13C NMR (63 MHz, D2O) δ (ppm) 175.2, 162.9 (J 242), 140.6
(J 7), 134.6, 130.6 (J 8), 127.2, 124.5 (J 3), 115.4 (J 21), 115.1
(J 21), 80.0, 72.7, 69.6, 67.1, and 38.2; υmax (KBr)/cm-1 3400,
1716, and 1592; MS (CI) m/z (%) 265 (MH+ - H2O); HRMS
calcd for C14H14O4F (MH+ - H2O) 265.0876, found 265.0870.
(1R,3R,4R)-1-(2′-F lu or oben zyloxy)-3,4-d ih yd r oxycyclo-
h ex-5-en -1-ca r boxylic a cid (4d ): 70% overall; retention time
28 min; [R]D -5° (c 0.7 in H2O); 19F NMR (282 MHz, D2O) δ
25
(ppm) -117.0 (dt, 1F, J 10.5 and 6.3); 1H NMR (250 MHz, D2O)
δ (ppm) 7.41-7.29 (m, 2H), 7.15-7.02 (m, 2H), 5.94 (d, 1H, J
10.1), 5.86 (dd, 1H, J 10.1 and 1.7), 4.50 (s, 2H), 4.02 (dt, 1H,
J 8.2 and 1.7), 3.78 (ddd, 1H, J 12.1, 8.2, and 3.6), 2.18 (ddd,
1H, J 13.7, 3.6, and 1.4), and 1.90 (dd, 1H, J 13.7 and 12.1);
13C NMR (63 MHz, D2O) δ (ppm) 177.8, 161.2 (J 244), 135.0,
131.8 (J 4), 130.9 (J 8), 127.0, 124.6 (J 18), 124.7, 115.7 (J
21), 80.3, 72.7, 69.6, 61.6 (J 4), and 37.8 (CH2); υmax (KBr)/
cm-1 3420, 1717, and 1589; MS (CI) m/z (%) 265 (MH+ - H2O);
HRMS calcd for C14H14O4F (MH+ - H2O) 265.0876, found
265.0876.
(1R ,3R ,4R )-3,4-Dih yd r oxy-1-(4′-ca r b oxy)b e n zyloxy-
cycloh ex-5-en -1-ca r boxylic a cid (4e): 65% overall; reten-
tion time 27.5 min; mp 161-162 °C; [R]D +16° (c 1.3 in
CH3OH); H NMR (250 MHz, CD3OD) δ (ppm) 7.94 (d, 2H, J
25
Gen er a l P r oced u r e for th e P r ep a r a tion of Resin s
10a -g a n d 20a -g. To preswollen resin in dry DMF at 0 °C
and under inert atmosphere was added sodium hydride (6
equiv, ca. 60% in mineral oil). The resultant suspension was
shook for 1 h at room temperature, and then the corresponding
bromide (10 equiv) and 15-crown-5 ether (0.3 equiv) were
added. The resin was gently stirred and heated between 40
and 80 °C for 24-48 h. The resin was thoroughly washed with
THF, THF-10% HCl (3:1), THF, and dry DCM to afford after
drying in vacuo resins 10a -i and 20a -g.
1
8.2), 7.45 (d, 2H, J 8.2), 6.01 (d, 1H, J 10.1), 5.87 (dd, 1H, J
10.1 and 1.9), 4.61 (d, 1H, J 11.8), 4.54 (d, 1H, J 11.8), 3.99-
3.82 (m, 2H), 2.26 (dd, 1H, J 13.2 and 3.4), and 1.96 (dd, 1H,
J 13.2 and 11.5); 13C NMR (63 MHz, CD3OD) δ (ppm) 177.0,
170.0, 145.6, 135.7, 130.6, 128.5, 128.2, 80.7, 74.1, 70.9, 67.5,
and 30.8; υmax (KBr)/cm-1 3444 and 1697; MS (CI) m/z (%) 291
(MH+
H2O); HRMS calcd for C15H15O6 (MH+ - H2O)
-
291.0869, found 291.0873.
Gen er a l Clea va ge P r oced u r e. The resin was treated with
(1:1) TFA-DCM (1 mL per 100 mg of resin) at room temper-
ature for 1 h. The resin was filtered and washed with dry
DCM. The filtrate was evaporated and dried in vacuo. The
obtained residue was redissolved in THF and treated with 5
equiv of 0.5 M aqueous lithium hydroxide. The mixture was
stirred for 30 min and then it was diluted with water and
washed with diethyl ether (3×). The aqueous extract was
treated with Amberlite IR-120 (H+) until pH 6. The resin was
filtered and washed with water. The filtrate was lyophilized
to afford a colorless oil or a foam unless specified.
(1R,3R,4R)-3,4-Dih yd r oxy-1-(4′-h yd r oxym eth yl)ben zyl-
oxycycloh ex-5-en -1-ca r b oxylic a cid (4f): 75% overall;
retention time 21 min; mp 143-144 °C; [R]25 +21° (c 0.8 in
1
CH3OH); H NMR (250 MHz, CD3OD) δ (ppm) 7.30 (d, 2H, J
8.3), 7.24 (d, 2H, J 8.3), 5.97 (d, 1H, J 10.1), 5.85 (dd, 1H, J
10.1 and 1.9), 4.52 (s, 2H), 4.47 (d, 1H, J 11.0), 4.42 (d, 1H, J
11.0), 3.93 (dt, 1H, J 7.9 and 1.9), 3.83 (ddd, 1H, J 11.6, 7.9,
and 3.5), 2.22 (m, 1H), and 1.90 (dd, 1H, J 13.3 and 11.6); 13
C
NMR (63 MHz, CD3OD) δ (ppm) 176.7, 141.9, 139.0, 135.9,
129.1, 127.9, 127.8, 80.4, 74.1, 70.8, 68.0, 65.0, and 39.6; υmax
(KBr)/cm-1 3372 and 1699; MS (CI) m/z (%) 259 (MH+ - 2H2O);
HRMS calcd for C15H15O4 (MH+ - 2H2O) 259.0970, found
259.0967.
(1R,3R,4R)-1-Ben zyloxy-3,4-d ih yd r oxycycloh ex-5-en -1-
ca r boxylic a cid (4a ): 85% overall; retention time 25 min;

Inhibitors of S. coelicolor Type II Dehydroquinase
J ournal of Medicinal Chemistry, 2003, Vol. 46, No. 26 5741
(1R,3R,4R)-3,4-Dih yd r oxy-1-(4′-cya n o)ben zyloxycyclo-
h ex-5-en -1-ca r boxylic a cid (4g): 70% overall; retention time
δ (ppm) 8.24 (br d, 2H, J 8.9), 7.69 (br d, 2H, J 8.9), 6.00 (s,
2H), 4.73 (d, 1H, J 12.6), 4.67 (d, 1H, J 12.6), 4.39 (br s, 1H),
4.26 (br s, 1H), 4.03 (m, 1H), 3.91 (m, 1H), 3.77 (s, 3H), 2.30
(ddd, 1H, J 13.5, 3.6, and 0.9), and 1.98 (dd, 1H, J 13.5 and
11.7); 13C NMR (62.5 MHz, CD3OD) δ (ppm) 172.1, 146.9,
146.3, 136.3, 127.7, 124.4, 122.9, 78.8, 72.8, 69.1, 65.4, 51.5,
and 38.0; υmax (NaCl)/cm-1 3390, 1740, and 1606; MS (CI) m/z
33 min; mp 81-82 °C; [R]D +18° (c 0.6 in CH3OH); 1H NMR
25
(250 MHz, CD3OD) δ (ppm) 7.66 (d, 2H, J 8.3), 7.55 (d, 2H, J
8.3), 6.01 (d, 1H, J 10.1), 5.87 (dd, 1H, J 10.1 and 2.0), 4.64
(d, 1H, J 12.3), 4.56 (d, 1H, J 12.3), 3.98 (dt, 1H, J 7.8, 2.0,
and 1.8), 3.86 (ddd, 1H, J 11.5, 7.8, and 3.6), 2.26 (ddd, 1H, J
13.3, 3.6, and 2.2), and 1.98 (dd, 1H, J 13.3 and 11.5); 13C NMR
(63 MHz, CD3OD) δ (ppm) 177.2, 146.4, 135.7, 133.1, 129.3,
128.3, 119.8, 111.9, 80.8, 74.1, 70.9, 67.1, and 39.7; υmax (KBr)/
cm-1 3410, 2232, and 1716; MS (CI) m/z (%) 272 (MH+ - H2O);
HRMS calcd for C15H14NO4 (MH+ - H2O) 272.0923, found
272.0930.
(%) 306 (MH+ - H2O); HRMS calcd for C15H16NO6 (MH+
-
H2O) 306.0978, found 306.0978.
(1R,3R,4R)-3,4-Dih yd r oxy-1-(4′-n itr oben zyloxy)cyclo-
h ex-5-en -1-ca r boxylic Acid (4h ). A solution of the methyl
ester 15 (25 mg, 0.08 mmol) in 0.5 mL of THF and 0.4 mL of
0.5 M aqueous lithium hydroxide was stirred at room tem-
perature for 1 h. The resultant solution was diluted with water
and treated with Amberlite IR-120 (H+) until pH 6. The resin
was filtered and washed with water. The filtrate was concen-
trated to afford acid 4h (23 mg, 96%) as a pale yellow oil:
(1R,3R,4R)-4-Ben zoyloxy-1-h yd r oxycycloh ex-5-en -1,3-
ca r bola cton e (11). To a stirred solution of the silyl ether 6
(2.16 g, 5.78 mmol), under argon and at 0 °C, in 80 mL of a
dry THF, was added tetrabutylammonium fluoride (6.4 mL,
6.36 mmol, ca. 1.0 M in THF). After stirring for 30 min, dilute
HCl was added and the organic layer was extracted with DCM
(3×). The combined organic extracts were dried (anhydrous
Na2SO4) and concentrated under reduced pressure. The crude
reaction was purified by flash cromatography eluting with 75%
diethyl ether-hexanes and recrystallized from hexane to yield
retention time 40 min; [R]D +13° (c 2.2 in CH3OH); 1H NMR
25
(250 MHz, CD3OD) δ (ppm) 8.38 (d, 2H, J 8.7), 7.82 (d, 2H, J
8.7), 6.23 (d, 1H, J 10.0), 6.09 (d, 1H, J 10.0 and 1.8), 4.90 (d,
1H, J 12.6), 4.82 (d, 1H, J 12.6), 4.20 (br d, 1H, J 7.8), 4.07
(m, 1H), 2.48 (br d, 1H, J 12.8), and 2.20 (dd, 1H, J 12.8 and
11.8); 13C NMR (63 MHz, CD3OD) δ (ppm) 177.5, 148.8, 148.6,
136.1, 129.5, 128.5, 124.5, 81.2, 74.3, 71.2, 67.2, and 40.0; υmax
(KBr)/cm-1 3419, 1718, and 1604; MS (CI) m/z (%) 292 (MH+
- H2O); HRMS calcd for C14H14NO6 (MH+ - H2O) 292.0821,
found 292.0821.
25
the alcohol 11 as white needles (1.38 g, 92%): [R]D -103° (c
1
0.6 in CHCl3); mp 104-105 °C (hexane); H NMR (250 MHz,
CDCl3) δ (ppm) 8.03 (dd, 2H, J 8.5 and 1.4), 7.60 (m, 1H), 7.46
(t, 1H, J 7.5), 6.30 (d, 1H, J 9.7), 5.88 (ddd, 1H, J 9.7, 3.3, and
1.1), 5.54 (t, 1H, J 3.0), 4.90 (m, 1H), 3.79 (br s, 1H), 2.55 (ddd,
1H, J 11.7, 5.2, and 1.7), and 2.49 (d, 1H, J 11.7); 13C NMR
(63 MHz, CDCl3) δ (ppm) 177.1, 165.2, 138.6, 133.7, 129.8,
128.9, 128.6, 124.2, 74.3, 73.3, 66.1, and 37.2; υmax (KBr)/cm-1
3478, 1775, and 1722; MS (CI) m/z (%) 261 (MH)+; HRMS calcd
for C14H13O5 (MH+) 261.0763, found 261.0755. Anal. Calcd for
(1R,3R,4R)-4-Ben zoyloxy-1-(2′-n itr oben zyloxy)cycloh ex-
5-en -1,3-ca r b ola ct on e (13). To a stirred solution of the
alcohol 11 (200 mg, 0.77 mmol) in dry DMF (7 mL) at 0 °C
under inert atmosphere was added sodium hydride (37 mg,
0.92 mmol, ca. 60% in mineral oil). After 30 min at this
temperature 2-nitrobenzyl bromide (216 mg, 1.00 mmol),
tetrabutylammonium iodide (28 mg, 0.08 mmol), and 15-
crown-5 ether (20 µL, 0.08 mmol) were added. The resultant
blue suspension was heated at 80 °C for 24 h. The suspension
was diluted successively with diethyl ether and water. The
organic layer was separated and the aqueous layer was
extracted with diethyl ether (3 × 20 mL). All the combined
organic layers were dried (anhydrous Na2SO4), filtered, and
evaporated. The crude residue was purified by flash chroma-
tography eluting with 70% DCM-hexanes to afford 196 mg
(64%) of ether 13 as amorphous beige solid and also 24 mg of
C
14H12O5: C, 64.60; H, 4.65. Found: C, 64.60; H, 4.65.
(1R,3R,4R)-4-Ben zoyloxy-1-(4′-n itr oben zyloxy)cycloh ex-
5-en -1,3-ca r b ola ct on e (12). To a stirred solution of the
alcohol 11 (93 mg, 0.36 mmol) in dry DMF (1.6 mL) at 0 °C
under inert atmosphere was added sodium hydride (17 mg,
0.43 mmol, ca. 60% in mineral oil). After 30 min at this
temperature 4-nitrobenzyl bromide (156 mg, 0.72 mmol),
tetrabutylammonium iodide (27 mg, 0.07 mmol), and 15-
crown-5 ether (10 µL, 0.04 mmol) were added. The resultant
blue suspension was heated at 80 °C for 24 h. The suspension
was diluted successively with diethyl ether and water. The
organic layer was separated and the aqueous layer was
extracted with diethyl ether (3 × 20 mL). All the combined
organic layers were dried (anhydrous Na2SO4), filtered, and
evaporated. The crude residue was purified by flash chroma-
tography eluting with 70% DCM-hexanes to afford 58 mg
(41%) of ether 12 as an amorphous pale yellow solid and also
28 mg (30%) of starting material was recovered: mp 135-136
°C (diethyl ether); [R]D25 -254° (c 0.9 in CHCl3); 1H NMR (250
MHz, CDCl3) δ (ppm) 8.23 (dt, 2H, J 8.8 and 2.1), 8.04 (dt,
2H, J 7.0, and 1.5), 7.60 (m, 3H), 7.46 (m, 2H), 6.41 (dd, 1H,
J 9.8 and 2.0), 5.98 (ddd, 1H, J 9.8, 3.4, and 2.1), 5.57 (t, 1H,
J 3.0), 4.93 (m, 1H), 4.84 (d, 1H, J 12.6), 4.79 (d, 1H, J 12.6),
2.71 (ddd, 1H, J 10.9, 6.0, and 2.0), and 2.40 (d, 1H, J 10.9);
13C NMR (63 MHz, CDCl3) δ (ppm) 173.1, 165.1, 147.6, 144.5,
136.2, 133.8, 129.8, 128.9, 128.6, 127.8, 125.0, 123.7, 78.9, 73.5,
66.4, 66.3, and 33.9; υmax (KBr)/cm-1 1801, 1722, and 1604;
MS (CI) m/z (%) 396 (MH+); HRMS calcd for C21H18NO7 (MH+)
396.1083, found 396.1066.
25
starting material (12%) was recovered: [R]D -290° (c 1.5 in
CHCl3); 1H NMR (250 MHz, CDCl3) δ (ppm) 8.05 (m, 4H), 7.70
(t, 1H, J 7.6), 7.61 (t, 1H, J 7.4), 7.47 (t, 3H, J 7.5), 6.43 (dd,
1H, J 9.8 and 1.5), 5.98 (ddd, 1H, J 9.8, 3.3, and 1.3), 5.57 (t,
1H, J 3.1), 5.17 (d, 1H, J 14.3), 5.01 (d, 1H, J 14.3), 4.94 (m,
1H), 2.74 (ddd, 1H, J 10.9, 6.1, and 1.9), and 2.50 (d, 1H, J
10.9); 13C NMR (63 MHz, CDCl3) δ (ppm) 173.0, 165.1, 146.7,
136.3, 134.0, 133.7, 129.8, 128.9, 128.9, 128.6, 128.4, 125.3,
124.7, 78.7, 73.6, 66.2, 64.3, and 33.8; υmax (NaCl)/cm-1 1793
and 1718; MS (CI) m/z (%) 396 (MH+); HRMS calcd for
C
21H18NO7 (MH+) 396.1083, found 396.1083.
(1R,3R,4R)-4-Hyd r oxy-1-(2′-n itr oben zyloxy)cycloh ex-
5-en -1,3-ca r bola cton e (16) a n d Meth yl (1R,3R,4R)-3,4-
Dih yd r oxy-1-(2′-n itr oben zyloxy)cycloh ex-5-en -1-ca r box-
yla te (17). To a stirred suspention of the benzoate 13 (127
mg, 0.32 mmol) in dry methanol (3 mL) was added potassium
cyanide (24 mg, 0.37 mmol). The resultant solution was stirred
at room temperature for 1 h. DCM was added and then water.
The organic layer was separated and the aqueous layer was
extracted twice with DCM. All the combined organic extracts
were dried (anhydrous Na2SO4), filtered, and evaporated. The
obtained residue was purified by flash chromatography eluting
with diethyl ether-ethyl acetate (90:10) to yield carbolactone
16 (24 mg, 26%) and diol 17 (65 mg, 63%).
Meth yl (1R,3R,4R)-3,4-Dih ydr oxy-1-(4′-n itr oben zyloxy)-
cycloh ex-5-en -1-ca r boxyla te (15). To a stirred suspension
of the benzoate 12 (62 mg, 0.16 mmol) in dry methanol (1.5
mL) was added potassium cyanide (12 mg, 0.18 mmol). The
resultant red solution was stirred at room temperature for 1
h. DCM was added and then water. The organic layer was
separated and the aqueous layer was extracted twice with
DCM. All the combined organic extracts were dried (anhydrous
Na2SO4), filtered, and evaporated. The obtained residue was
purified by flash chromatography eluting with 40% ethyl
acetate-hexanes to yield diol 15 (31 mg, 60%) as a pale yellow
Data for carbolactone 16: [R]D -175° (c 1.4 in CHCl3); 1H
25
NMR (250 MHz, CDCl3) δ (ppm) 8.10 (dd, 1H, J 8.2 and 1.3),
7.96 (dd, 1H, J 7.9 and 1.0), 7.68 (m, 1H), 7.47 (m, 1H), 6.25
(dd, 1H, J 9.8 and 1.6), 5.87 (ddd, 1H, J 9.8, 3.3, and 1.2), 5.11
(d, 1H, J 14.4), 4.95 (d, 1H, J 14.4), 4.74 (m, 1H), 4.33 (t, 1H,
J 3.1), 2.61 (ddd, 1H, J 10.9, 6.1, and 2.1), and 2.40 (d, 1H, J
10.9); 13C NMR (63 MHz, CDCl3) δ (ppm) 173.8, 146.7, 134.4,
oil: [R]D +5° (c 0.9 in CH3OH); 1H NMR (250 MHz, CD3OD)
25

5742 J ournal of Medicinal Chemistry, 2003, Vol. 46, No. 26
Gonza´lez-Bello et al.
134.0, 133.8, 128.9, 128.4, 128.3, 124.8, 79.1, 76.2, 65.1, 64.2,
and 32.8; υmax (NaCl)/cm-1 3459 and 1790; MS (CI) m/z (%)
292 (MH+); HRMS calcd for C14H14NO6 (MH+) 292.0821, found
292.0827.
162.8 (J 242), 133.5, 131.0 (J 8), 130.9, 128.9, 115.7 (J 21),
80.0, 74.2, 71.3, 67.8, and 39.7; υmax (Nujol)/cm-1 3391, 3333,
and 1713; MS (CI) m/z (%) 265 (MH+ - H2O); HRMS calcd for
C
14H14O4F (MH+ - H2O) 265.0876, found 265.0875.
Data for diol 17: [R]D25 +43° (c 1.5 in CHCl3); 1H NMR (250
MHz, CDCl3) (ppm) δ 7.97 (dd, 1H, J 8.2 and 1.0), 7.79 (dd,
1H, J 7.8 and 1.0), 7.60 (dt, 1H, J 7.6 and 1.0), 7.38 (dt, 1H, J
7.8 and 1.0), 5.95 (s, 2H), 4.84 (d, 1H, J 14.5), 4.78 (d, 1H, J
14.5), 4.08 (d, 1H, J 8.0), 3.93 (m, 1H), 3.72 (s, 3H), 2.33 (dd,
1H, J 13.6 and 3.1), and 1.98 (dd, 1H, J 13.6 and 12.1); 13C
NMR (63 MHz, CDCl3) δ (ppm) 172.4, 147.1, 135.8, 134.2,
133.7, 129.3, 128.1, 125.1, 124.4, 78.8, 73.2, 69.8, 63.8, 52.7,
and 38.0; υmax (NaCl)/cm-1 3390 and 1739; MS (CI) m/z (%)
(1R,3R,4R)-4-(3′-F lu or oben zyloxy)-1,3-d ih yd r oxycyclo-
h ex-5-en -1-ca r boxylic a cid (5c): 75% overall; retention time
31 min; [R]25 -127° (c 0.9 in H2O); 19F NMR (282 MHz, D2O)
D
δ (ppm) -111.6 (dt, 1F, J 9.5 and 5.6); 1H NMR (250 MHz,
D2O) δ (ppm) 7.21 (m, 1H), 7.04 (m, 2H), 6.92 (m, 1H), 5.79 (d,
1H, J 10.0), 5.61 (d, 1H, J 10.0), 4.55 (s, 2H), 3.86 (m, 2H),
and 1.94 (d, 2H, J 6.0); 13C NMR (63 MHz, D2O) δ (ppm) 178.1,
162.9 (J 245), 140.3 (J 7), 131.1, 130.6 (J 8), 128.5, 124.4 (J
3), 115.3 (J 22), 115.2 (J 21), 80.2, 73.9, 71.2, 67.7, and 39.5;
υmax (NaCl)/cm-1 3410 and 1726; MS (CI) m/z (%) 283 (MH+);
HRMS calcd for C14H16O5F (MH+) 283.0982, found 283.0978.
292 (MH+ - HOCH3); HRMS calcd for C14H14NO6 (MH+
-
HOCH3) 292.0821, found 292.0823.
(1R,3R,4R)-3,4-Dih yd r oxy-1-(2′-n itr oben zyloxy)cyclo-
h ex-5-en -1-ca r boxylic Acid (4i). A solution of the methyl
ester 17 (65 mg, 0.20 mmol) in 2 mL of THF and 1 mL of 0.5
M aqueous lithium hydroxide was stirred at room temperature
for 1 h. The resultant solution was diluted with water and
treated with Amberlite IR-120 (H+) until pH 6. The resin was
filtered and washed with water. The filtrate was concentrated
to afford acid 4i (48 mg, 78%) as a beige oil: retention time
(1R,3R,4R)-4-(2′-F lu or oben zyloxy)-1,3-d ih yd r oxycyclo-
h ex-2-en -1-ca r boxylic a cid (5d ): 70% overall; retention time
29 min; [R]25 -134° (c 3.0 in H2O); 19F NMR (282 MHz, D2O)
D
1
δ (ppm) -116.9 (dt, 1F, J 10.5 and 6.4); H NMR (250 MHz,
D2O) δ (ppm) 7.26 (t, 1H, J 7.2), 7.13 (m, 1H), 6.98 (t, 1H, J
7.0), 6.90 (t, 1H, J 8.9), 5.70 (d, 1H, J 9.9), 5.55 (d, 1H, J 9.9),
4.55 (s, 2H), 3.83 (m, 2H), and 1.92 (m, 2H); 13C NMR (63 MHz,
D2O) δ (ppm) 178.2, 161.2 (J 245), 131.5 (J 4), 130.9, 130.8 (J
10), 128.7, 124.7 (J 3), 124.5 (J 15), 115.7 (J 21), 80.4, 73.9,
67.7, 65.9 (J 3), and 39.5; υmax (Nujol)/cm-1 3430 and 1718;
MS (CI) m/z (%) 283 (MH+); HRMS calcd for C14H16O5F (MH+)
283.0982, found 283.0970.
35 min; [R]D -19° (c 0.7 in CH3OH); 1H NMR (250 MHz,
25
CD3OD) δ (ppm) 7.96 (dd, 1H, J 8.2 and 1.1), 7.87 (br d, 1H,
J 7.9), 7.63 (m, 1H), 7.42 (m, 1H), 5.97 (d, 1H, J 10.1), 5.86 (d,
1H, J 10.1), 4.84 (s, 2H), 3.94 (d, 1H, J 7.8), 3.85 (m, 1H), 2.26
(br d, 1H, J 13.6), and 1.96 (dd, 1H, J 13.6 and 12.0); 13C NMR
(63 MHz, CD3OH) δ (ppm) 178.4, 148.6, 136.7, 135.1, 134.5,
131.5, 128.9, 125.2, 73.6, 73.2, 71.2, 64.4, and 39.7; υmax (KBr)/
cm-1 3398, 1707, and 1578; MS (CI) m/z (%) 292 (MH+ - H2O);
HRMS calcd for C14H14NO6 (MH+ - H2O) 292.0821, found
292.0820.
P r ep a r a t ion of R esin 18. To a stirred solution of the
alcohol 11 (1.43 g, 5.48 mmol) in dry DMF (25 mL) at 0 °C
under inert atmosphere was added sodium hydride (264 mg,
6.60 mmol, ca. 60% in mineral oil). After 30 min at this
temperature the resultant suspension was added via cannula
to a preswollen PS-brominated Wang resin (1.4 g, ∼1.6 mmol/
g, ∼2.24 mmol) in dry DMF (16 mL) at 0 °C and under inert
atmosphere. Then, 15-crown-5 ether (60 µL, 0.32 mmol) was
added and the resultant suspension was shook at 0 °C for 30
min and at 80 °C for 48 h. The resin was thoroughly washed
with DMF, DMF-water (3:1), THF, and dry DCM and dried
under high vacuum to afford brown beads of 11 (1.4 g): gel-
phase FTIR (cm-1) 1794, 1718, and 1611; gel-phase 13C NMR
(63 MHz, CDCl3) δ (ppm) 137.0, 133.6, 129.5, 114.6, 73.6, 70.0,
67.5, 40.4, and 34.0.
(1R ,3R ,4R )-4-(4′-Ca r b oxyb e n zyloxy)-1,3-d ih yd r oxy-
cycloh ex-5-en -1-ca r boxylic a cid (5e): 68% overall; reten-
tion time 32 min; mp 187-188 °C; [R]25 -152° (c 1.1 in
D
1
CH3OH); H NMR (300 MHz, CD3OD) δ (ppm) 7.98 (d, 2H, J
7.8), 7.50 (d, 2H, J 7.8), 5.90 (dd, 1H, J 1.8 and 9.9), 5.71 (d,
1H, J 9.9), 4.80 (s, 2H), 4.04 (m, 1H), 3.92 (d, 1H, J 7.5), and
2.10 (m, 2H); 13C NMR (75 MHz, CD3OD) δ (ppm) 177.7, 168.9,
144.6, 130.0, 130.0, 129.8, 127.5, 80.5, 73.6, 71.0, 68.2, and
39.9; υmax (KBr)/cm-1 3423 and 1699; MS (CI) m/z (%) 291
(MH+ - H2O); HRMS calcd for C15H14O6 (MH+ - H2O)
291.0869, found 291.0880.
(1R,3R,4R)-1-Hyd r oxy-4-(4′-h yd r oxym eth yl)ben zyloxy-
cycloh ex-5-en -1-ca r boxylic a cid (5f): 77% overall; retention
time 21 min; [R]25 -136° (c 1.3 in CH3OH); 1H NMR (250
D
MHz, CD3OD) δ (ppm) 7.42 (d, 2H, J 8.1), 7.36 (d, 2H, J 8.1),
5.90 (dd, 1H, J 10.0 and 1.7), 5.72 (d, 1H, J 10.0), 4.75 (s, 2H),
4.62 (s, 2H), 4.06 (m, 1H), 3.93 (d, 1H, J 7.4), and 2.11 (m,
2H); 13C NMR (63 MHz, CD3OD) δ (ppm) 178.6, 142.1, 138.9,
131.2, 130.7, 129.1, 128.0, 81.0, 74.6, 72.6, 69.2, 64.9, and 40.0;
υ
max (NaCl)/cm-1 3364 and 1730; MS (CI) m/z (%) 277 (MH+
-
H2O); HRMS calcd for C15H17O5 (MH+ - H2O) 277.1076, found
P r ep a r a tion of Resin 19. To preswollen resin 18 (1.1 g,
∼1.3 mmol) in dry THF (6 mL) was added a solution of
potassium cyanide (136 mg, 2.09 mmol) in methanol (2 mL).
The resultant suspension was bubbled through with argon for
2 h at room temperature. The resin was thoroughly washed
with THF, methanol, THF, and dry DCM to afford after drying
in vacuo pale brown beads of resin 19 (∼1 g): gel-phase FTIR
(cm-1) 3392, 1792, 1732, and 1604; gel-phase 13C NMR (63
MHz, CDCl3) δ (ppm) 129.6, 114.6, 70.2, 67.4, 53.4, and 40.4.
277.1083.
(1R,3R,4R)-1,3-Dih yd r oxy-4-(4′-cya n oben zyloxy)cyclo-
h ex-5-en -1-ca r boxylic a cid (5g): 70% overall; retention time
37 min; mp 137-138 °C; [R]25 -157° (c 1.0 in CH3OH); 1H
D
NMR (300 MHz, D2O) δ (ppm) 7.40 (d, 2H, J 7.8), 7.28 (d, 2H,
J 7.8), 5.72 (d, 1H, J 9.9), 5.57 (d, 1H, J 9.9), 4.55 (s, 2H), 3.84
(m, 2H), and 1.92 (d, 2H, J 7.2); 13C NMR (75 MHz, D2O) δ
(ppm) 178.3, 143.6, 132.6, 130.4, 129.0, 128.5, 119.6, 110.4,
80.4, 73.9, 70.7, 67.8, and 39.6; υmax (KBr)/cm-1 3398, 2231,
and 1720; MS (CI) m/z (%) 272 (MH+ - H2O); HRMS calcd for
(1R,3R,4R)-4-Ben zyloxy-1,3-d ih yd r oxycycloh ex-5-en -1-
ca r boxylic a cid (5a ): 75% overall; retention time 27 min;
C
15H14O4N (MH+ - H2O) 272.0923, found 272.0923.
[R]D -128° (c 3.4 in H2O); 1H NMR (250 MHz, D2O) δ (ppm)
25
(1R,3R,4R)-1-(ter t-Bu tyldim eth ylsilyloxy)-4-(4′-n itr oben -
7.15-7.03 (m, 5H), 5.65 (dd, 1H, J 9.8 and 1.6), 5.41 (d, 1H, J
9.8), 4.44 (d, 1H, J 13.8), 4.39 (d, 1H, J 13.8), 3.77-3.60 (m,
2H), and 1.74 (d, 2H, J 6.2); 13C NMR (63 MHz, CD3OD) δ
(ppm) 177.8, 139.0, 131.3, 130.6, 128.9, 129.0, 128.6, 81.0, 75.0,
72.7, 69.1, and 40.8; υmax (KBr)/cm-1 3420 and 1706; MS
(+FAB) m/z (%) 247 (MH+ - H2O); HRMS calcd for C14H15O4
(MH+ - H2O) 247.0970, found 247.0964.
zyloxy)cycloh ex-5-en -1,3-ca r bola cton e (21). To a stirred
solution of the alcohol 7 (164 mg, 0.61 mmol) in dry DMF (6
mL) at 0 °C under inert atmosphere was added sodium hydride
(29 mg, 0.73 mmol, ca. 60% in mineral oil). After 30 min at
this temperature 4-nitrobenzyl bromide (171 mg, 0.79 mmol),
tetrabutylammonium iodide (23 mg, 0.06 mmol), and 15-
crown-5 ether (10 µL, 0.08 mmol) were added. The resultant
blue suspension was heated at 80 °C for 48 h. The suspension
was diluted successively with diethyl ether and water. The
organic layer was separated and the aqueous layer was
extracted with diethyl ether (3 × 20 mL). All the combined
organic layers were dried (anhydrous Na2SO4), filtered, and
evaporated. The crude residue was purified by flash chroma-
tography eluting with 70% DCM-hexanes to afford 60 mg
(1R,3R,4R)-4-(4′-F lu or oben zyloxy)-1,3-d ih yd r oxycyclo-
h ex-5-en -1-ca r boxylic a cid (5b): 72% overall; retention time
28 min; [R]25 -133° (c 0.9 in H2O); 19F NMR (282 MHz, D2O)
D
δ (ppm) -112.5 (tt, 1F, J 9.2 and 5.2); 1H NMR (250 MHz,
D2O) δ (ppm) 7.35 (t, 2H, J 5.8), 7.04 (t, 2H, J 8.3), 5.87 (d,
1H, J 9.7), 5.65 (d, 1H, J 9.7), 4.61 (s, 2H), 3.92 (m, 2H), and
1.98 (d, 2H, J 6.8); 13C NMR (63 MHz, D2O) δ (ppm) 179.0,

Inhibitors of S. coelicolor Type II Dehydroquinase
J ournal of Medicinal Chemistry, 2003, Vol. 46, No. 26 5743
(24%) of ether 21 as amorphous pale yellow solid: [R]D25 -172°
(1R,3R,4R)-1-Hyd r oxy-4-(2′-n itr oben zyloxy)cycloh ex-
5-en -1,3-ca r bola cton e (24). To a stirred solution of the silyl
ether 22 (165 mg, 0.41 mmol), under argon and at 0 °C, in 4
mL of a dry THF, was added tetrabutylammonium fluoride
(0.45 mL, 0.45 mmol, ca. 1.0 M in THF). After stirring for 30
min, dilute HCl was added and the organic layer was extracted
with DCM (3×). The combined organic extracts were dried
(anhydrous Na2SO4) and concentrated under reduced pressure.
The crude reaction was purified by flash cromatography
eluting with 75% diethyl ether-hexanes to yield the alcohol
24 as beige oil (85 mg, 71%): [R]D25 -269° (c 0.9 in CHCl3); 1H
NMR (250 MHz, CDCl3) δ (ppm) 8.04 (dd, 1H, J 8.1 and 1.2),
7.74-7.61 (m, 2H), 7.46 (dt, 1H, J 8.1 and 1.8), 6.16 (ddd, 1H,
J 9.8, 1.5, and 0.7), 5.84 (ddd, 1H, J 9.8, 3.4, and 1.1), 5.08 (d,
1H, J 14.0), 5.00 (d, 1H, J 14.0), 4.85 (m, 1H), 4.05 (dt, 1H, J
3.4 and 0.7), 3.81 (br s, 1H), 2.46 (ddd, 1H, J 11.0, 5.4, and
1.5) and 2.39 (d, 1H, J 11.0); 13C NMR (63 MHz, CDCl3) δ
(ppm) 177.5, 147.3, 137.0, 133.7, 133.6, 128.9, 128.6, 125.1,
124.8, 74.3, 73.4, 72.4, 69.0, and 36.8; υmax (NaCl)/cm-1 3432
and 1790; MS (CI) m/z (%) 292 (MH+); HRMS calcd for
1
(c 2.0 in CHCl3); H NMR (250 MHz, CDCl3) δ (ppm) 8.21 (d,
2H, J 8.8), 7.50 (d, 2H, J 8.8), 6.15 (ddd, 1H, J 9.8, 1.7, and
1.0), 5.78 (ddd, 1H, J 9.8, 3.2, and 1.1), 4.77 (s, 2H), 4.73 (m,
1H), 3.98 (t, 1H, J 3.2), 2.42 (ddd, 1H, J 10.6, 5.2, and 1.8),
2.37 (d, 1H, J 10.6), 0.90 (s, 9H), 0.17 (s, 3H), and 0.14 (s, 3H);
13C NMR (63 MHz, CDCl3) δ (ppm) 175.3, 147.5, 144.9, 138.9,
127.7, 124.2, 123.7, 75.0, 73.4, 72.4, 70.9, 37.7, 25.5, 17.9, and
-3.1; υmax (NaCl)/cm-1 1793; MS (CI) m/z (%) 406 (MH+);
HRMS calcd for
406.1676.
C
20H28O6NSi (MH+) 406.1686, found
(1R,3R,4R)-1-Hyd r oxy-4-(4′-n itr oben zyloxy)cycloh ex-
5-en -1,3-ca r bola cton e (23). To a stirred solution of the silyl
ether 21 (95 mg, 0.23 mmol), under argon and at 0 °C, in 2
mL of dry THF, was added tetrabutylammonium fluoride (0.25
mL, 0.25 mmol, ca. 1.0 M in THF). After stirring for 30 min,
dilute HCl was added and the organic layer was extracted with
DCM (3×). The combined organic extracts were dried (anhy-
drous Na2SO4) and concentrated under reduced pressure. The
crude reaction product was purified by flash cromatography
eluting with diethyl ether to afford alcohol 23 as a pale yellow
C
14H14O6N (MH+) 292.0821, found 292.0824.
oil (29 mg, 43%): [R]D -265.5° (c 1.2, in CHCl3); 1H NMR
25
(1R,3R,4R)-1,3-Dih yd r oxy-4-(2′-n itr oben zyloxy)cyclo-
h ex-5-en -1-ca r b oxylic Acid (5i). A solution of the carbo-
lactone 24 (23 mg, 0.08 mmol) in 1 mL of THF and 0.4 mL of
0.5 M aqueous lithium hydroxide was stirred at room tem-
perature for 1 h. The resultant solution was diluted with water
and treated with Amberlite IR-120 (H+) until pH 6. The resin
was filtered and washed with water. The filtrate was concen-
trated to afford acid 5h (23 mg, 94%) as a beige amorphous
(250 MHz, CDCl3) δ (ppm) 8.22 (d, 2H, J 8.7), 7.51 (d, 2H, J
8.7), 6.17 (d, 1H, J 9.8), 5.82 (ddd, 1H, J 9.8, 3.3, and 1.0),
4.81 (m, 1H), 4.78 (s, 2H), 4.02 (t, 1H, J 3.3), 3.35 (br s, 1H),
and 2.44 (m, 2H); 13C NMR (63 MHz, CDCl3) δ (ppm) 177.2,
147.6, 144.8, 137.4, 127.7, 125.1, 123.8, 74.5, 73.4, 72.2, 71.0,
and 36.9; υmax (NaCl)/cm-1 3405 and 1784; MS (CI) m/z (%)
292 (MH+); HRMS calcd for C14H14O6N (MH+) 292.0821, found
292.0826.
25
solid: retention time 46 min; [R]D -59° (c 1.0, in CH3OH);
1H NMR (250 MHz, CD3OD) δ 8.01 (dd, 1H, J 8.1 and 1.2),
7.89 (br d, 1H, J 8.4), 7.68 (dt, 1H, J 7.6 and 1.2), 7.49 (m,
1H), 5.89 (d, 1H, J 9.7), 5.72 (d, 1H, J 9.7), 5.09 (br s, 2H),
4.03 (m, 1H), 3.93 (d, 1H, J 7.5), and 2.05 (m, 2H); 13C NMR
(63 MHz, CD3OD) δ 176.7, 148.1, 135.0, 133.5, 130.4, 129.6,
129.3, 128.4, 124.4, 81.3, 73.6, 68.7, 68.4, and 40.1; υmax (NaCl)/
cm-1 3397, 1716, and 1609; MS (CI) m/z (%) 292 (MH+ - H2O);
HRMS calcd for C14H14O6N (MH+ - H2O): 292.0821, found
292.0819.
(1R,3R,4R)-1,3-Dih yd r oxy-4-(4′-n itr oben zyloxy)cyclo-
h ex-5-en -1-ca r boxylic Acid (5h ). A solution of the carbo-
lactone 23 (28 mg, 0.10 mmol) in 1 mL of THF and 0.5 mL of
0.5 M aqueous lithium hydroxide was stirred at room tem-
perature for 1 h. The resultant solution was diluted with water
and treated with Amberlite IR-120 until pH 6. The resin was
filtered and washed with water. The filtrate was concentrated
to afford acid 5h (23 mg, 74%) as a pale yellow amorphous
25
solid: retention time 48 min; [R]D -121° (c 1.1, in CH3OH);
1H NMR (250 MHz, CD3OD) δ (ppm) 8.20 (d, 2H, J 8.8), 7.65
(d, 2H, J 8.8), 5.93 (dd, 1H, J 10.0 and 1.8), 5.73 (d, 1H, J
10.0), 4.84 (s, 2H), 4.04 (m, 1H), 3.93 (dt, 1H, J 7.9 and 1.8),
and 2.07 (m, 2H); 13C NMR (63 MHz, CD3OD) δ (ppm) 176.8,
147.6, 147.1, 130.4, 129.3, 128.1, 123.4, 81.1, 73.5, 70.4, 68.2,
and 40.2; υmax (KBr)/cm-1 3528 and 1596; MS (CI) m/z (%) 310
(MH+); HRMS calcd for C14H16NO7 (MH+) 310.0928, found
310.0928.
Ack n ow led gm en t . Financial support from the
Xunta de Galicia (PGIDIT02RAG20901PR) and from
the Spanish Ministry of Science and Technology
(SAF2001-3120) is gratefully acknowledged. E.L. thanks
the Spanish Ministry of Education and Culture for an
FPU scholarship. M.D.T. thanks the Fundac¸ao para a
Cieˆncia e a Tecnologia for a scholarship.
(1R,3R,4R)-1-(ter t-Bu tyldim eth ylsilyloxy)-4-(2′-n itr oben -
zyloxy)cycloh ex-5-en -1,3-ca r bola cton e (22). To a stirred
solution of the alcohol 7 (138 mg, 0.51 mmol) in dry DMF (4
mL) at 0 °C under inert atmosphere was added sodium hydride
(27 mg, 0.66 mmol, ca. 60% in mineral oil). After 30 min at
this temperature 2-nitrobenzyl bromide (165 mg, 0.77 mmol),
tetrabutylammonium fluoride (28 mg, 0.08 mmol), and 15-
crown-5 ether (10 µL, 0.08 mmol) were added. The resultant
blue suspension was heated at 80 °C for 24 h. The suspension
was diluted successively with diethyl ether and water. The
organic layer was separated and the aqueous layer was
extracted with diethyl ether (3 × 20 mL). All the combined
organic layers were dried (anhydrous Na2SO4), filtered, and
evaporated. The crude residue was purified by flash chroma-
tography eluting with 25% diethyl ether-hexanes to afford
1
Su p p or tin g In for m a tion Ava ila ble: Copies of H NMR
and 13C NMR spectra of compounds 4a -i and 5a -i, 19F NMR
spectra of compounds 4b-d and 5b-d , and gel-phase 13C NMR
spectra of 8, 9, 18, and 19. This material is available free of
charge via the Internet at http://pubs.acs.org.
Refer en ces
(1) Abell, C. Enzymology and Molecular Biology of the Shikimate
Pathway. In Comprehensive Natural Products Chemistry, ed.
Sankawa, U., Pergamon, Elsevier Science Ltd.: Oxford, 1999;
p 573.
(2) Roberts, F.; Roberts, C. W.; J ohnson, J . J .; Kyle, D. E.; Krell,
T.; Coggins, J . R.; Coombs, G. H.; Milhous, W. K.; Tzipori, S.;
Ferguson, D. J . P.; Chakrabarti, D.; McLeod, R. Evidence for
the shikimate pathway in apicomplexan parasites. Nature 1998,
393, 801.
(3) (a) Steinrucken, H. C.; Amerhein, N. 5-Enolpyruvylshikimate-
3-phosphate synthase of Klebsiella pneumoniae 2. Inhibition by
glyphosate [N-(phosphonomethyl)glycine]. Eur. J . Biochem.
1984, 143, 351. (b) Steinrucken, H. C.; Amerhein, N. The
herbicide glyphosate is a potent inhibitor of 5-enolpyruvyl-
shikimic acid 3-phosphate synthase. Biochem. Biophys. Res.
Commun. 1980, 94, 1207.
(4) Chauduri, C.; Ducan, K.; Graham, L. D.; Coggins, J . R. Identi-
fication of the active-site lysine residues of two biosynthetic
3-dehydroquinases. Biochem. J . 1990, 275, 1.
25
111 mg (54%) of ether 22 as amorphous beige solid: [R]D
1
-205° (c 1.2 in CHCl3); H NMR (250 MHz, CDCl3) δ (ppm)
8.06 (dd, 1H, J 8.1 and 1.0), 7.67 (m, 2H), 7.48 (dt, 1H, J 8.2
and 1.8), 6.15 (dd, 1H, J 9.8 and 1.7), 5.81 (ddd, 1H, J 9.8,
3.3, and 1.9), 5.08 (d, 1H, J 13.9), 5.00 (d, 1H, J 13.9), 4.78
(m, 1H), 4.02 (t, 1H, J 3.1), 2.43 (ddd, 1H, J 10.8, 5.8, and
1.9), 2.33 (d, 1H, J 10.8), 0.91 (s, 9H), 0.18 (s, 3H), and 0.14
(s, 3H); 13C NMR (63 MHz, CDCl3) δ (ppm) 175.4, 147.4, 138.7,
133.6, 129.0, 128.6, 124.7, 124.3, 75.0, 73.2, 72.5, 69.0, 37.7,
25.5, 17.9, and -3.2; υmax (NaCl)/cm-1 1798; MS (CI) m/z (%)
406 (MH+); HRMS calcd for C20H28O6NSi (MH+) 406.1686,
found 406.1703.
(5) White, P. J .; Young, J .; Hunter, I. S.; Nimmo, H. G.; Coggins, J .
R. The purification and characterization of 3-dehydroquinase
from Streptomyces coelicolor. Biochem J . 1990, 265, 735.

5744 J ournal of Medicinal Chemistry, 2003, Vol. 46, No. 26
Gonza´lez-Bello et al.
(6) (a) Shneier, A.; Kleanthous, C.; Deka, R.; Coggins, J . R.; Abell,
C. Observation of an imine intermediate on dehydroquinase by
electrospray mass spectrometry. J . Am. Chem. Soc. 1991, 113,
9416. (b) Leech, A. P.; J ames, R.; Coggins, J . R.; Kleanthous, C.
Mutagenesis of active site residues in type I dehydroquinase
from Escherichia coli. Stalled catalysis in a histidine to alanine
mutant. J . Biol. Chem. 1995, 270, 25827. (c) Hanson, K. R.; Rose,
I. A. The absolute stereochemical course of citric acid biosyn-
thesis. Proc. Natl. Acad. Sci. U.S.A. 1963, 50, 981. (d) Smith, B.
W.; Turner, M. J .; Haslam, E. Stereochemical course of the
3-dehydroquinate dehydratase reaction and a novel preparation
of shikimic acid labeled with isotopic hydrogen at C-2. J . Chem.
Soc., Chem. Commun. 1970, 842. (e) Haslam, E.; Turner, M. J .;
Sargent, D.; Thompson, R. S. Shikimate pathway. I. Preparation
of stereospecifically labeled 2-deuterio derivatives of 3-de-
hydroquinic acid. J . Chem. Soc. (C) 1971, 1489. (f) Harris, J .;
Kleanthous, C.; Coggins, J . R.; Hawkins, A. R.; Abell, C.
Different mechanistic and sterechemical courses for the reactions
catalysed by type I and type II dehydroquinases. J . Chem. Soc.,
Chem. Commun. 1993, 13, 1080. (g) Butler, J . R.; Alworth, W.
L.; Nugent, M. Mechanism of dehydroquinase catalysed dehy-
dratation. I. Formation of a schiff base intermediate. J . Am.
Chem. Soc. 1974, 96, 1617. (h) Chaudhuri, S.; Duncan, K.;
Graham,. L. D.; Coggins, J . R. Identification of the active-site
lysine residues of two biosynthetic 3-dehydroquinases. Biochem-
istry 1991, 275, 1.
(7) Harris, J .; Gonza´lez-Bello, C.; Kleanthous, C.; Coggins, J . R.;
Hawkins, A. R.; Abell, C. Evidence from kinetic isotope studies
for an enolate intermediate in the mechanism of type II
dehydroquinases. Biochem. J . 1996, 319, 333.
(8) Gonza´lez-Bello, C.; Harris, J . M.; Manthey, M. K.; Coggins, J .
R.; Abell, C. Irreversible Inhibition of Type I dehydroquinase
by substrates for type II dehydroquinases. Bioorg. Med. Chem.
Lett. 2000, 10, 407.
(9) Fredrickson, M.; Parker, E. J .; Hawkins, A. R.; Coggins, J . R.;
Abell, C. Selective inhibition of type II dehydroquinases. J . Org.
Chem. 1999, 64, 2612.
(10) Roszak, A. W.; Robinson, D. A.; Krell, T.; Hunter, I. S.; Freder-
ickson, M.; Abell, C.; Coggins, J . R.; Lapthorn, A. The structure
and mechanism of the type II dehydroquinase from Steptomyces
Coelicolor. Structure 2002, 10, 493.
(12) Gonza´lez, C.; Carballido, M.; Castedo, L. Synthesis of poly-
hydroxycyclohexanes from (-)-quinic acid. J . Org. Chem. 2003,
68, 2248.
(13) Brominated Wang resin was prepared from Wang resin as
indicated previously Lee, C. E.; Kick, E. K.; Ellman, J . A. General
solid-phase synthesis approach to prepare mechanism-
based aspartyl protease inhibitor libraries. Identification of
potent Cathepsin D inhibitors. J . Am. Chem. Soc. 1998, 120,
9735.
(14) The loading of the resin was calculated by treatment of a small
amount of resin (50 mg) with in 1 mL of TFA/DCM (1:1) for 1 h.
The resin was filtered and washed with THF. The filtrate was
evaporated. The obtained residue was redissolved in DCM and
washed with a diluted solution of sodium bicarbonate. The
organic layer was dried (anhydrous Na2SO4), filtered, and
evaporated.
(15) 4-(tert-Butyldimethylsilyloxymethyl)benzyl bromide was pre-
pared from methyl 4-bromomethylbenzoate by reduction with
DIBAL-H at -78 °C in THF followed by protection of the
obtained alcohol with TBSOTf/Et3N in DCM at 0 °C.
(16) (a) J ones, G.; Willett, P.; Glen, R. C. Molecular recognition of a
receptor sites using a genetic alorithm with a description of
desolvation. J . Mol. Biol. 1995, 245, 43. (b) J ones, G.; Willett,
P.; Glen, R. C.; Leach, A. R.; Taylor, R. Development and
validation of a genetic alorithm for flexible docking. J . Mol. Biol.
1997, 267, 727.
(17) Frisch, M. J .; Trucks, G. W.; Schlegel, H. B.; Scuseria, G. E.;
Robb, M. A.; Cheeseman, J . R.; Zakrzewski, V. G.; J . A.
Montgomery, J .; Stratmann, R. E.; Burant, J . C.; Pprich, S.;
Millam, J . M.; Daniels, A. D.; Kudin, K. N.; Strain, M. C.; Farkas,
O.; Tomasi, J .; Barone, V.; Cossi, M.; Cammi, R.; Mennucci, B.;
Pomelli, C.; Adamo, C.; Clifford, S.; Ochterski, J .; Petersson, G.
A.; Ayala, P. Y.; Cui, Q.; Morokuma, K.; Malick, D. K.; Rabuck,
A. D.; Raghavachari, K.; Foresman, J . B.; Cioslowski, J .; Ortiz,
J . V.; Baboul, A. G.; Stefanov, B. B.; Liu, G.; Liashenko, A.;
Piskorz, P.; Komaromi, I.; Gomperts, R.; Martin, R. L.; Fox, D.
J .; Keith, T.; Al-Laham, M. A.; Peng, C. Y.; Nanayakkara, A.;
Gonza´lez, C.; Challacombe, M.; Gill, P. M. W.; J ohnson, B. G.;
Chen, W.; Wong, M. W.; Andres, J . L.; Head-Gordon, M.;
Replogle, E. S.; Pople, J . A. Gaussian 98W (Revision A.9);
Gaussian, Inc., Pittsburgh, PA, 1998.
(11) Toscano, M. D.; Frederickson, M.; Evans, D. P.; Coggins, J . R.;
Abell, C.; Gonza´lez-Bello, C. Design, synthesis and evaluation
of bifunctional inhibitors of type II dehydroquinase. Org. Biomol.
Chem. 2003, 1, 2075.
J M030987Q