Crystal structure and total synthesis of globomycin: Establishment of relative and absolute configurations [5]

Full text of DOI:10.1021/ja002547j

10214
J. Am. Chem. Soc. 2000, 122, 10214-10215
Crystal Structure and Total Synthesis of
Globomycin: Establishment of Relative and Absolute
Configurations
Hiroshi Kogen,*,† Toshihiro Kiho,† Mizuka Nakayama,†
Youji Furukawa,‡ Takeshi Kinoshita,‡ and Masatoshi Inukai§
Exploratory Chemistry Research Laboratories
Biomedical Research Laboratories
and Lead DiscoVery Research Laboratories
Sankyo Co., Ltd., 2-58, Hiromachi 1-chome Shinagawa-ku
Tokyo 140-8710, Japan
Figure 1.
ReceiVed July 17, 2000
Globomycin (1) was isolated as a cyclic depsipeptide antibiotic
against Gram-negative bacteria,1,2 and as the first natural product1c,3
which contains both L-allo-Ile and L-allo-Thr, in 1978. 1 has been
proven to be a specific inhibitor of signal peptidase II (prolipo-
protein signal peptidase),4 which processes the acylated precursor
form of lipoproteins into apolipoprotein and signal peptide in
Escherichia coli.5 A breakthrough in lipoprotein research was the
finding that signal peptidase II activity is specifically inhibited
by 1.5,6 To our knowledge, 1 is the only specific inhibitor of signal
peptidase II known for the present. 1 causes the accumulation of
the acylated forms of lipoprotein in the cytoplasmic membrane
and, consequently, death of the cell.7a Therefore, signal peptidase
II represents an attractive target for developing a new class of
antibiotics that works with a different mechanism from currently
available drugs. Globomycin (1) has been used routinely to
demonstrate the acylation of newly identified lipoprotein,8 and
since then it has been widely used for controlling the maturation
of the lipopeptides.7 Although 1 has been an invaluable tool in
studies of lipoprotein biosynthesis9 to date, its structure remained
obscure for almost two decades. An initial structure elucidation
of 1 was only able to determine the absolute stereochemistry of
the L-allo-Thr-L-Ser-L-allo-Ile moiety. However, the relative and
the absolute stereochemistry of the 3-hydroxy-2-methylnonanoyl-
Figure 2. ORTEP drawing of globomycin (1).
N-Me-Leu moiety still remained ambiguous.1c,2b,10,11 Herein we
wish to report the complete stereochemical structure of 1 from
the X-ray diffraction analysis and asymmetric total synthesis
of 1.
The structure of 1, which was unequivocally determined by
X-ray crystallography, revealed the following two points: the
relative stereochemistry of 1 is as shown in Figure 2, and all the
amides are in the trans conformation.12 The crystal structure of 1
also suggested that the carbonyl oxygen of the L-allo-Ile forms
an intramolecular hydrogen bond with the NH of glycine (the
O17-N31 distance is 2.8 Å).
To develop the synthetic route to various globomycin analogues
in search of a more effective inhibitor of signal peptidase II, and
to investigate an SAR study of its interesting inhibition activity,
we explored a total synthesis of 1. Our synthetic route to 1 consists
of (i) an asymmetric synthesis of (2R,3R)-3-hydroxy-2-methyl-
nonanoic acid (2), (ii) convergent coupling of three components
(2, 7, and 11), and (iii) macrolactamization (Scheme 1).
Synthesis of (+)-2 was achieved by an anti-selective asym-
metric boron-mediated aldol reaction which was reported by
Masamune.13 O-Benzyl-L-serine allyl ester (3) was condensed with
N-Boc-L-allo-isoleucine (4) in the presence of the 1H-benzotria-
zol-1-yloxytripyrrolidinophosphonium hexafluorophosphate (Py-
BOP)14 reagent to provide dipeptide 5 in 97% yield. The Boc
group was subsequently removed by using 4 N HCl in ethyl
acetate, and the resulting residue was coupled with N-Boc-N-
† Exploratory Chemistry Research Laboratories.
‡ Biomedical Research Laboratories.
§ Lead Discovery Research Laboratories.
(1) (a) Inukai, M.; Enokita, R.; Torikata, A.; Nakahara, M.; Iwado, S.; Arai,
M. J. Antibiot. 1978, 31, 410-420. (b) Inukai, M.; Nakajima, M.; Osawa,
M.; Haneishi, T.; Arai, M. J. Antibiot. 1978, 31, 421-425. (c) Nakajima, M.;
Inukai, M.; Haneishi, T.; Terahara, A.; Arai, M.; Kinoshita, T.; Tamura, C. J.
Antibiot. 1978, 31, 426-432.
(2) Isolation of globomycin and its analogue has been reported by Omoto
et al. also following ref 1c. (a) Omoto, S.; Suzuki, H.; Inouye, S. J. Antibiot.
1979, 32, 83-86. (b) Omoto, S.; Ogino, H.; Inouye, S. J. Antibiot. 1981, 34,
1416-1423.
(3) Bycroft, B. W. In Amino acids, Peptides, and Proteins; Sheppard, R.
C., Senior Reporter; Specialist Periodical Report; The Chemical Society:
London, 1979; Vol. 12, Chaptor 4.
(4) Three types of signal peptidase, LepB (leader peptidase), signal peptidase
II, and type IV prepilin peptidase, have been identified. Pugsley, A. P.
Microbiol. ReV. 1993, 57, 50-108.
(5) (a) Inukai, M.; Takeuchi, M.; Shimizu, K.; Arai, M. J. Antibiot. 1978,
31, 1203-1205. (b) Hussain, M.; Ichihara, S.; Mizushima, S. J. Biol. Chem.
1980, 255, 3707-3712. (c) Dev, I. K.; Harvey, R. J.; Ray, P. H. J. Biol.
Chem. 1985, 260, 5891-5894.
(6) (a) Ichihara, S.; Hussain, M.; Mizushima, S. J. Biol. Chem. 1981, 256,
3125-3129. (b) Witke, C.; Go¨tz, F. FEMS Microl. Lett. 1995, 126, 233-
240.
(7) Recent work in this area: (a) Cavard, D. Arch. Microbiol. 1998, 171,
50-58. (b) Ochsner, U. A.; Vasil, A. I.; Johnson, Z.; Vasil, M. L. J. Bacteriol.
1999, 181, 1099-1109. (c) Paitan, Y.; Orr, E.; Ron, E. Z.; Rosenberg, E. J.
Bacteriol. 1999, 181, 5644-5651. (d) Tibor, A.; Decelle, B.; Letesson, J.-J.
Infect. Immun. 1999, 67, 4960-4962. For review, see: Cavard, D.; Oudega,
B. In Bacteriocins, Microcins and Lantibiotics; Lazdunski, C., Pattus, F., Eds.;
NATO ASI series; Springer: Berlin, Heidelberg, New York, 1992; Vol. 65,
pp 297-305.
(8) For review, see: Braun, B.; Wu, H. C. In Bacterial Cell Wall; Ghuysen,
J. M., Hakenbeck, R., Eds.; Elsevier: Amsterdam, 1994; Chapter 14.
(9) Sakka, K.; Watanabe, T.; Beers, R.; Wu, H. C. J. Bacteriol. 1987, 169,
3400-3408.
(10) Omoto et al. presumed that the relative stereochemistry of 3-hydroxy-
1
2-methylnonanoic acid is anti by H NMR. See ref 2b.
(11) Partial racemization of N-Me-Leu easily occurred during acid hy-
drolysis. See refs 1c and 2b.
(12) X-ray data for 1: C32H57N5O9, colorless prisms, hexagonal, P61, a )
b ) 26.711(3) Å, c ) 9.884(4) Å, R ) â ) 90.0000°, V ) 6107(2) Å3, Z )
6, dcalc ) 1.070 g/cm3, R ) 0.053, Rw ) 0.070, GOF ) 1.16.
(13) Abiko, A.; Liu, J. F.; Masamune, S. J. Am. Chem. Soc. 1997, 119,
2586-2587. For synthetic detail of (+)-2, see the Supporting Information.
(14) Coste, J.; Le-Nguyen, D.; Castro, B. Tetrahedron Lett. 1990, 31, 205-
208.
10.1021/ja002547j CCC: $19.00 © 2000 American Chemical Society
Published on Web 09/27/2000

Communications to the Editor
J. Am. Chem. Soc., Vol. 122, No. 41, 2000 10215
Scheme 1a
a Conditions: (a) PyBOP, Et3N, 0 °C, CH2Cl2 (97%); (b) (i) 4 N HCl-AcOEt, room temperature; (ii) PyBOP, 6, Et3N, CH2Cl2, 0 °C (2 steps, 95%);
(c) (i) 4 N HCl-AcOEt, room temperature; (ii) (+)-2, DEPC, Et3N, THF, 0 °C (2 steps, 95%); (d) (i) 10, PyBop, CH2Cl2, 0 °C to room temperature
(93%); (ii) Pd/C-H2, room temperautre (100%); (e) 11, DMAP, DCC, cat. CSA, CH2Cl2, 0 °C to room temperature (96%); (f) (i) n-Bu4NF, AcOH,
THF, 0 °C to room temperature (99%); (ii) cat. Pd(PPh3)4, morpholine, THF, room temperature (97%); (iii) TFA, CH2Cl2, room temperature; (iv)
HATU, i-Pr2NEt, THF, 0 °C to room temperature (2 steps, 45%); (v) Pd(OH)2, room temperature (96%).
Me-L-leucine (6) under PyBOP conditions to afford tripeptide 7
in 95% yield. Deprotection of the Boc group in 7, followed by
diethyl phosphorocyanidate (DEPC)-induced amide bond forma-
tion with the (2R,3R)-3-hydroxy-2-methylnonanoic acid (2),
completed the formation of N-acyl tripeptide 8 in 95% yield. The
dipeptide fragment 11 was prepared by a condensation between
N-Boc-O-TBS-L-allo-threonine (9) and glycine benzyl ester (10)
with the PyBOP method, followed by hydrogenolysis over 10%
Pd/C to give dipeptide 11 in 93% yield. The esterification of the
tripeptide 8 with the dipeptide unit 11 using DCC and DMAP15
in the presence of a catalytic amount of 10-camphorsulfonic acid
(CSA) afforded the protected depsipeptide 12 in 96% yield.
Removal of the TBS group using TBAF (99%) was followed by
deprotection of the allyl group with Pd(PPh3)4 (97%) and
N-terminal Boc with TFA to afford the amine as its salt which
was then set for macrolactamization. This intermediate underwent
a cyclization in the presence of HATU16 and N,N′-diisopropyl-
ethylamine yielding O-benzyl globomycin (45% in two steps).
Finally, removal of the benzyl group by hydrogenolysis afforded
Figure 3. Conformation of the N-methyl amide moiety with selected
NOESY correlations in CD3OD.19
isomers are derived from the rotation of the acyl N-methyl amide
moiety. The correlation among the N-methyl group, C3-H, and
C19-H in the major isomer indicated the trans conformation for
the amide moiety which is a similar result to that of the X-ray
analysis. However, the correlation between C3-H and C19-H
in the minor isomer suggests that it adopted the cis-form in the
amide moiety (Figure 3). Synthetic 1 against Escherichia coli
ATCC 11303 showed the same degree of antimicrobial activity
(MIC ) 0.2 µg/mL) as that initially observed for natural
globomycin.1b However, the antimicrobial activity of the O-benzyl
globomycin was extremely diminished. This result suggests that
the serine hydroxyl group of globomycin is quite essential for
the antimicrobial activity.
In summary, the relative and absolute configurations of
globomycin (1) were determined by X-ray analysis and an
asymmetric total synthesis which was achieved by convergent
coupling of three components followed by macrolactamization.
Further synthetic investigations for SAR study are currently
underway.
globomycin (1) in 96% yield [mp 114-116 °C, [R]24 +23.8°
D
(c 0.50, MeOH)]. All physical properties [mixed mp, 500 MHz
1H (16 mM) and 125 MHz 13C (18 mM) NMR, [R]D, and IR] of
synthetic 1 are identical with those of the authentic sample.17 Thus,
the correlation of synthetic and natural 1 establishes the absolute
configuration as shown in Figure 1.
1
The H NMR spectrum suggests that 1 exists as a mixture of
two rotational isomers in solution. The ratio is dependent on the
solvent (major/minor18 ) 6:1 in CDCl3, 3:1 in CD3OD at 23 °C).
The examination of NOESY experiments suggests that these
(15) Boden, E. P.; Keck, G. E. J. Org. Chem. 1985, 50, 2394-2395.
(16) (a) Carpino, L. A.; El-Faham, A. J. Org. Chem. 1994, 59, 695. (b)
Carpino, L. A. J. Am. Chem. Soc. 1993, 115, 4397. (c) Carpino, L. A.; El-
Faham, A. J. Org. Chem. 1995, 60, 3561. For a review of peptide coupling
methods, see: Humphrey, J. M.; Chamberlin, R. Chem. ReV. 1997, 97, 2243-
2266.
Acknowledgment. We thank Dr. S. Miyakoshi (Sankyo Co., Ltd.)
for the antimicrobacterial assay of globomycin.
Supporting Information Available: X-ray structural information and
stereoview of 1, experimental procedures, and characterization data for
(17) Natural globomycin: mp 115-116 °C (from CH3CN), [R]23D +24.1°
(c 0.50, MeOH). For isolation and purification of natural 1, see ref 1b.
(18) The isomer ratio was determined by 1H NMR analysis of N-Me
protons.
1
all new compounds, and H and 13C spectra of natural and synthetic 1
(PDF). This material is available free of charge via the Internet at
http://pubs.acs.org.
(19) The same NOESY correlations for major isomer were measured in
CDCl3.
JA002547J