Total synthesis of mycocyclosin

Article abstract of DOI:10.1021/ol300831t

The first total synthesis of mycocyclosin, a diketopiperazine natural product isolated from M. tuberculosis, is described. While direct oxidative coupling of tyrosine phenolic groups was unsuccessful, construction of the highly strained bicyclic framework was successfully accomplished through an intramolecular Miyaura-Suzuki cross-coupling to generate the biaryl linkage.

Full text of DOI:10.1021/ol300831t

ORGANIC
LETTERS
2012
Vol. 14, No. 9
2402–2405
Total Synthesis of Mycocyclosin
James R. Cochrane, Jonathan M. White, Uta Wille, and Craig A. Hutton*
School of Chemistry and Bio21 Molecular Science and Biotechnology Institute,
University of Melbourne, Parkville, VIC 3010, Australia
chutton@unimelb.edu.au
Received April 2, 2012
ABSTRACT
The first total synthesis of mycocyclosin, a diketopiperazine natural product isolated from M. tuberculosis, is described. While direct oxidative
coupling of tyrosine phenolic groups was unsuccessful, construction of the highly strained bicyclic framework was successfully accomplished
through an intramolecular MiyauraꢀSuzuki cross-coupling to generate the biaryl linkage.
Mycocyclosin 1 (Figure 1) is a diketopiperazine second-
ary metabolite produced by Mycobacterium tuberculosis,
discovered in 2009 by Belin and co-workers.1 The bio-
synthesis of mycocyclosin proceeds through initial con-
densation of two molecules of tyrosyl-tRNA 2 to generate
cyclo(L-TyrꢀL-Tyr) 3 (Scheme 1). Formation of the dike-
topiperazine 3 is catalyzed by the enzyme cyclodityrosine
synthetase. This enzyme is the first characterized member
of a recently discovered family of enzymes known as cyclo-
dipeptide synthetases (CDPS) that structurally resemble
the catalytic domain of class Ic tRNA synthetases.2ꢀ5
Subsequent oxidative coupling of the tyrosine aromatic
rings of 3 is catalyzed by the cytochrome P450 enzyme
CYP121, which generates a 3,30-dityrosine-type biaryl
cross-link to complete the bicyclic framework of 1.1ꢀ3
Recently, the gene encoding the CYP121 P450 enzyme,
rv2276, was shown to be essential for M. tuberculosis
viability,6 leading to speculation that mycocyclosin has a
vital but as yet undetermined cellular role in M. tuberculosis
survival.
Furthermore, the M. tuberculosis CYP121 P450 enzyme
appears to be a target for the azole antifungal/antimyco-
bacterial drugs,6 highlighting the potential of this enzyme
as a novel M. tuberculosis drug target. The use of azole-
type therapeutics often leads to serious side effects due to
their nonselective inhibition of host P450 enzymes, and
thus the design of selective inhibitors of the CYP121 P450
enzyme, based on structural analogy to mycocyclosin,
could ultimately provide new therapeutics for tuberculosis
with fewer side effects.
(1) Belin, P.; Le Du, M. H.; Fielding, A.; Lequin, O.; Jacquet, M.;
Charbonnier, J.-B.; Lecoq, A.; Thai, R.; Courc-on, M.; Masson, C.;
Dugave, C.; Genet, R.; Pernodet, J.-L.; Gondry, M. Proc. Natl. Acad.
Sci. U.S.A. 2009, 106, 7426–7431.
(2) Gondry, M.; Sauguet, L.; Belin, P.; Thai, R.; Amouroux, R.;
Tellier, C.; Tuphile, K.; Jacquet, M.; Braud, S.; Courc-on, M.; Masson,
C.; Dubois, S.; Lautru, S.; Lecoq, A.; Hashimoto, S.-I.; Genet, R.;
Pernodet, J.-L. Nat. Chem. Biol. 2009, 5, 414–420.
(3) Vetting, M. W.; Hegde, S. S.; Blanchard, J. S. Nat. Chem. Biol.
2010, 6, 797–799.
(4) Sauguet, L.; Moutiez, M.; Li, Y.; Belin, P.; Seguin, J.; Le Du,
M. H.; Thai, R.; Masson, C.; Fonvielle, M.; Pernodet, J. L.; Charbonnier,
J. B.; Gondry, M. Nucleic Acids Res. 2011, 39, 4475–4489.
(5) Bonnefond, L.; Arai, T.; Sakaguchi, Y.; Suzuki, T.; Ishitani, R.;
Nureki, O. Proc. Natl. Acad. Sci. U.S.A. 2011, 108, 3912–3917.
Figure 1. Structures of mycocyclosin 1 and related natural
products herqulines A and B.
r
10.1021/ol300831t
Published on Web 04/20/2012
2012 American Chemical Society

Construction of a molecular model of mycocyclosin 1
suggests the bicyclic compound is significantly strained. In
this regard, mycocyclosin is reminiscent of strained cyclo-
phane-type natural products containing biaryl linkages,
such as haouamine and rhazinal, which have attracted
significant recent interest as synthetic targets.7ꢀ13 Myco-
cyclosin also bears close resemblance to the natural pro-
ducts herquline A and B (Figure 1), isolated by Omura and
co-workers from Penicillium herquei,14ꢀ16 and potentially
could be a biosynthetic precursor of such compounds.
The unusual structure of mycocyclosin, together with its
intriguing role in M. tuberculosis biology, prompted us to
investigate a route toward its total synthesis.
Boc-group and cyclization generated the diketopiperazine
3 in good yield as a single stereoisomer.
Scheme 2. Synthesis of L,L-Cyclodityrosine 3
Direct oxidative coupling of 3 was attempted using the
vanadium oxyfluoride-promoted process developed by
Evans and co-workers.20 However, under these conditions
only dimers and trimers of the diketopiperazine 3 were
detected, with no evidence of intramolecular phenolic
coupling to generate mycocyclosin 1. In retrospect, direct
oxidative coupling to generate strained biaryl linkages has
met with only limited success.10,13
Scheme 1. Biosynthesis of Mycocyclosin 1
Scheme 3. Synthesis of Diketopiperazine 11
In pursuit of a synthetic route to mycocyclosin 1, we
envisaged a biomimetic approach through initial diketo-
piperazine synthesis followed by oxidative phenolic cou-
pling. Cyclodityrosine has been prepared by condensation
of L-tyrosine in ethylene glycol at high temperature;17
however, this method has been shown to cause epimeriza-
tion, resulting in mixtures of stereoisomers.18 Accordingly,
cyclo(L-TyrꢀL-Tyr) 3 was prepared by a stepwise method.19
Tyrosine was converted to the protected dipeptide 5
through standard amino acid protecting group and pep-
tide coupling techniques (Scheme 2). Removal of the
(6) McLean, K. J.; Carroll, P.; Lewis, D. G.; Dunford, A. J.; Seward,
H. E.; Neeli, R.; Cheesman, M. R.; Marsollier, L.; Douglas, P.; Smith,
W. E.; Rosenkrands, I.; Cole, S. T.; Leys, D.; Parish, T.; Munro, A. W.
J. Biol. Chem. 2008, 283, 33406–33416.
ꢀ
(7) Garrido, L.; Zubı
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68, 293–299.
a, E.; Ortega, M. J.; Salva, J. J. Org. Chem. 2003,
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(9) Burns, N. Z.; Krylova, I. N.; Hannoush, R. N.; Baran, P. S. J. Am.
Chem. Soc. 2009, 131, 9172–9173.
Attention was therefore turned to a metal-catalyzed
cross-coupling approach, which required the correspond-
ing iodotyrosine-containing diketopiperazine 10. Accord-
ingly, cyclo(L-TyrꢀL-Tyr) 3 was treated with iodine in the
presence of silver sulfate; however, to drive the iodination
of both tyrosine residues to completion, small amounts of
the corresponding tri-iodinated product were produced,
which was difficult to remove. A more efficient route to
iodinated 10 was therefore pursued from L-iodotyrosine 6.
Following an analogous procedure to that used for the
(10) Wipf, P.; Furegati, M. Org. Lett. 2006, 8, 1901–1904.
(11) Grundl, M. A.; Trauner, D. Org. Lett. 2006, 8, 23–25.
(12) Baran, P. S.; Burns, N. Z. J. Am. Chem. Soc. 2006, 128, 3908–
3909.
(13) Bowie, A. L.; Trauner, D. J. Org. Chem. 2009, 74, 1581–1586.
(14) Omura, S.; Hirano, A.; Iwai, Y.; Masuma, R. J. Antibiot. 1979,
32, 786–790.
(15) Furusaki, A.; Matsumoto, T.; Ogura, H.; Takayanagi, H.;
Hirano, A.; Omura, S. J. Chem. Soc., Chem. Commun. 1980, 698.
(16) Enomoto, Y.; Shiomi, K.; Hayashi, M.; Masuma, R.; Kawakubo,
T.; Tomosawa, K.; Iwai, Y.; Omura, S. J. Antibiot. 1996, 49, 50–53.
(17) Xia, H.; Suo, Z. Y.; Qiang, G. J.; Chang, C. J. J. Appl. Polym.
Sci. 2008, 110, 2168–2178.
(20) (a) Evans, D. A.; Dinsmore, C. J.; Evrard, D. A.; DeVries, K. M.
J. Am. Chem. Soc. 1993, 115, 6426–6421. For application to dityrosine,
see: (b) Reetz, M.; Merk, C.; Mehler, G. Chem. Commun. 1998, 2075–
2076.
(18) Cook, B.; Hill, R. R.; Jeffs, G. E. J. Chem. Soc., Perkin Trans. 1
1992, 1199.
(19) Jung, M. E.; Rohloff, J. C. J. Org. Chem. 1985, 50, 4909–4913.
Org. Lett., Vol. 14, No. 9, 2012
2403

Table 1. Optimization of Biaryl Coupling/Cyclizationa
13
yield
(%)
entry
R
(equiv)
catalyst
b
1
Bn
Bn
H
ꢀ
NiCl2(PCy3)2, Zn, PPh3
0
0
c
2
ꢀ
Ni(PPh3)4
3
1.2
1.2
1.2
1.2
1.2
1.2
1.0
0.6
Pd(dppf) CH2Cl2
0
3
4
Bn
Bn
Bn
Bn
Bn
Bn
Bn
Pd(dppf) CH2Cl2
33
20
19
5
3
5d
6
Pd(dppf) CH2Cl2
3
Pd(OAc)2, Sphose
7f
8g
9
Pd(dppf) CH2Cl2
3
Pd(dppf) CH2Cl2
26
42
27
3
Pd(dppf) CH2Cl2
3
10
Pd(dppf) CH2Cl2
3
Figure 2. (a) ORTEP diagram of bis(O-benzyl)mycocyclosin 12.
(b) Representation of crystal structure with perturbed bond
lengths and angles highlighted. (c) Representation of out-of-
plane bend of biaryl bond of 12. (d) View down biaryl bond
(C15ꢀC8) highlighting twist of aromatic rings and out-of-plane
bend of biaryl bond with respect to both aromatic rings.
a Conditions: 10 or 11, 0.005 M in DMSO, 13, 6 equiv K2CO3,
catalyst (20 mol %), 90 °C. b DMF solvent. c DMF solvent, 1.5 equiv
Ni(0). d 80 °C. e ligand/catalyst 3:1. f 1,4-Dioxane solvent. g Microwave
irradiation, 90 °C, 2 h.
preparation of L,L-cyclodityrosine 3, the L,L-cyclodi(iodo-
tyrosine) 10 was generated from 6 in three steps and 55%
yield (Scheme 3). Benzylation of 10 gave the protected
compound 11 in good yield.
was attempted by varying temperature (entry 5), ligand
(entry 6), solvent (entry 7), use of microwave irradiation
(entry 8), and stoichiometry (entries 9 and 10). All conditions
provided the protected mycocyclosin derivative 12 in low to
moderate yield, with an optimum 42% yield obtained.
Crystals of 12 suitable for X-ray crystallographic ana-
lysis were grown from DMSO. The crystal structure of 12
confirms that the [8.2.2] bicyclic framework is highly
strained (see Figure 2). The geometry of the dityrosine
biaryl linkage is significantly distorted: the biaryl bond is
longer than normal (1.51 A; cf. 1.49 A for an average biaryl
bond),24 and the associated bond angles deviate greatly
from the idealized value of 120° (e.g., C7ꢀC8ꢀC15 108.1°,
C9ꢀC8ꢀC15 134.4°). Notably, the C8ꢀC15 biaryl bond is
bent considerably out-of-plane with respect to the aromatic
rings, displaying R-angles of 22.3° and 16.3°. These distor-
tions are among the greatest deviations known for strained
cyclophane natural products and derivatives.7,10,25,26
Attempted Ni-catalyzed Ullmann-type reductive coupling
of bis-iodide 11 did not proceed (Table 1, entry 1),21 returning
a mixture of starting material and protodeiodinated material.
Accordingly, we switched to a one-pot Pd-catalyzed Miyaura
borylationꢀSuzuki coupling method we have previously
employed for the preparation of dityrosine derivatives.22
Zhu et al. also employed such an approach in an intramole-
cular manner to generate the biaryl linkage in a biphenomy-
cin analogue.23 Treatment of L,L-cyclodi(iodotyrosine) 10
with bispinacolatodiboron 13 in the presence of Pd(dppf)Cl2
gave none of the cross-coupled product, with proto-deiodi-
nated material recovered (entry 3). In contrast, treatment of
the benzyl-protected L,L-cyclodi(iodotyrosine) 11 under the
same conditions gave the cross-coupled adduct 12 in moderate
yield (entry 4). Optimization of the cross-coupling reaction
The high degree of strain present in the bicyclic com-
pound 12 suggests it is remarkable that the cross-coupling
proceeds in even moderate yield.27 We speculate that the
(21) For use of Ni(0)-promoted reductive cyclization in the synthesis
of alnusone, see: (a) Semmelhack, M. F.; Ryono, L. S. J. Am. Chem.
Soc. 1975, 97, 3873–3875. For attempts at intramolecular Ni- and Pd-
catalyzed couplings toward the biaryl-containing ring in vancomycin,
see: (b) Boger, D. L.; Miyazaki, S.; Kim, S. H.; Wu, J. H.; Castle, S. L.;
Loiseleur, O.; Jin, Q. J. Am. Chem. Soc. 1999, 121, 10004–10011.
(c) Nicolaou, K. C.; Li, H.; Boddy, C. N. C.; Ramanjulu, J. M.; Yue,
intermediate biaryl-palladium species represents
a
€
T.-Y.; Natarajan, S.; Chu, X.-J.; Brase, S.; Rubsam, F. Chem.;Eur. J.
1999, 5, 2584–2601.
(22) (a) Hutton, C. A.; Skaff, O. Tetrahedron Lett. 2003, 44, 4895–
4898. (b) Skaff, O.; Jolliffe, K. A.; Hutton, C. A. J. Org. Chem. 2005, 70,
7353–7363.
€
(24) Allen, F. H.; Watson, D. G.; Brammer, L.; Orpen, A. G.; Taylor,
R. In International Tables for Crystallography, Vol. C; Prince, E., Ed.;
Wiley-VCH: 2006; Chapter 9.5, pp 790ꢀ811.
(25) Cram, D.; Cram, J. M. Acc. Chem. Res. 1971, 4, 204–213.
(26) Lee, T. J.; Rice, J. E.; Allen, W. D.; Remington, R. B.; Schaefer,
H. F., III. Chem. Phys. 1988, 123, 1–25.
(23) Carbonnelle, A.-C.; Zhu, J. Org. Lett. 2000, 2, 3477–3480.
2404
Org. Lett., Vol. 14, No. 9, 2012

‘tethered’, low-strain system in which the tyrosine C3
positions are held in close proximity, with reductive elim-
ination then proceeding to yield the cross-linked product
12. Such an approach proved successful in Trauner’s
synthesis of rhazinal in which Pd-catalyzed cross-coupling
effected formation of a strained biaryl system, whereas
oxidative direct coupling failed.13
Scheme 4. Synthesis of Mycocyclosin 1
Figure 3. Calculated potential energy surface for the atropi-
somerization of mycocyclosin.
H-bond between the phenolic OH groups; without this
stabilization the barrier to rotation would be higher. The
barrier to rotation is sufficiently low that rapid intercon-
version of atropisomers would occur at rt. The lowest
With bis(O-benzyl)-protected mycocyclosin 12 in hand,
standard hydrogenolysis conditions were investigated to
effect deprotection. However, under such conditions, com-
plete decomposition of the strainedbicyclic compound was
observed. Fortunately, treatment of 12 with trifluoroacetic
acid (TFA) in the presence of pentamethylbenzene as a
cation scavenger effected clean benzyl group cleavage to
generate mycocyclosin 1 in good yield (Scheme 4). The 1H
and 13C NMR spectra of synthetic mycocyclosin closely
matched that reported for the natural product.1
energy conformation of mycocyclosin 1 is 5.8 kJ molꢀ1
3
more stable than its atropisomer and matches the config-
uration of the atropisomer observed in the solid state of 12.
Low temperature NMR studies of mycocyclosin are in
agreement with the calculations, showing that interconver-
sion of the atropisomers occurs rapidly at rt, with a coales-
cence temperature <190 K (see Supporting Information).
In conclusion, an efficient synthesis of the highly
strained meta-cyclophane-type diketopiperazine natural
product, mycocyclosin 1, has been developed employing
an intramolecular, one-pot borylationꢀSuzuki cross-cou-
pling to generate the biaryl linkage. A synthetic route to 1
should allow investigations into the biological role of
mycocyclosin in Mycobacterium tuberculosis viability and
also provides a synthetic route suitable for elaboration to
the preparation of analogues of mycocyclosin in the search
for potential antimycobacterial compounds.
Considering the strain exhibited in the benzyl-protected
mycocyclosin 12, the natural product would be expected to
possess similar strain. Though the structure of mycocyclo-
1
sin 1 has been determined by mass spectrometry and H
and 13C NMR spectroscopy, no confirmation of the three-
dimensional structure, including analysis of the configura-
tion about the biaryl chirality axis, has been reported. We
considered that the contraction of the angles about the
biaryl linkage could increase the barrier to rotation such
that atropisomers are possible. Accordingly, we calculated
the ground-state structure of mycocyclosin 1 and the
barrier to rotation about the biaryl axis.28
Acknowledgment. Financial support from the Austra-
lian Research Council (DP110100112) is acknowledged.
This work was supported by the National Computing
Infrastructure (NCI) facility. Dr. Ian Luck (University of
Sydney) is thanked for assistance with VT NMR experi-
ments. Assoc. Prof. Spencer Williams (University of
Melbourne) is thanked for bringing this compound to
our attention.
The calculated ground-state structure of mycocyclosin 1
is in close agreement with the crystal structure of the
benzyl-protected derivative 12; the bond angles around
the biaryl linkage are similarly distorted from ideal geo-
metry, and the biaryl bond is significantly bent out-of-
plane with respect to the aromatic rings (20.7° in 1; cf. 22.3°
in 12). The barrier to rotation between atropisomers was
calculated to be 39.6 kJ molꢀ1 (Figure 3), which is in the
0 3
range expected for a 1,1 -disubstituted biaryl.29 The transi-
tion state to atropisomerization is stabilized by a strong
Supporting Information Available. Full experimental
1
details, characterization data, and H and 13C NMR
spectra for all new compounds. This material is available
free of charge via the Internet at http://pubs.acs.org. The
authors declare no competing financial interest.
(27) A yield of 45% was obtained for a related intramolecular cross-
coupling with much less strain; see ref 23.
(28) Calculations were performed using the BHandHLYP hybrid
density functional in combination with the 6-31þG* basis set.
(29) Leroux, F. ChemBioChem 2004, 5, 644–649.
The authors declare no competing financial interest.
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