Solid-phase total synthesis of bogorol A: Stereocontrolled construction of thermodynamically unfavored (E)-2-amino-2-butenamide

Article abstract of DOI:10.1021/acs.orglett.5b00769

Bogorol A [(E)-1], a potent antibiotic against methicillin-resistant Staphylococcus aureus and vancomycin-resistant Enterococcus spp., possesses a thermodynamically unfavored (E)-2-amino-2-butenamide within its linear dodecapeptide sequence. The highly efficient total synthesis of natural (E)-isomer (E)-1 and its artificial (Z)-isomer (Z)-1 by employing a full solid-phase strategy is reported. The (E)- and (Z)-2-amino-2-butenamide moieties were stereoselectively constructed by applying traceless Staudinger ligation on the resin. Interestingly, (E)- and (Z)-1 showed comparable antimicrobial activity (MIC = 4 μg/mL).

Full text of DOI:10.1021/acs.orglett.5b00769

Letter
pubs.acs.org/OrgLett
Solid-Phase Total Synthesis of Bogorol A: Stereocontrolled
Construction of Thermodynamically Unfavored (E)‑2-Amino-2-
butenamide
Tomoya Yamashita, Takefumi Kuranaga, and Masayuki Inoue*
Graduate School of Pharmaceutical Sciences, The University of Tokyo, Hongo, Bunkyo-ku, Tokyo 113-0033, Japan
S
* Supporting Information
ABSTRACT: Bogorol A [(E)-1], a potent antibiotic against
methicillin-resistant Staphylococcus aureus and vancomycin-
resistant Enterococcus spp., possesses a thermodynamically
unfavored (E)-2-amino-2-butenamide within its linear dodec-
apeptide sequence. The highly efficient total synthesis of
natural (E)-isomer (E)-1 and its artificial (Z)-isomer (Z)-1 by
employing a full solid-phase strategy is reported. The (E)- and (Z)-2-amino-2-butenamide moieties were stereoselectively
constructed by applying traceless Staudinger ligation on the resin. Interestingly, (E)- and (Z)-1 showed comparable antimicrobial
activity (MIC = 4 μg/mL).
acterial infections increasingly evade standard treatments
as resistance to multiple antibiotics spreads through the
bond; consequently, the E/Z geometry of the olefin impacts the
dynamic behavior, overall three-dimensional structure, and
biological activity.7,8 The characteristic dehydroamino acid
residue of (E)-1, together with its important antibacterial
function, motivated us to undertake the total chemical
construction of both natural (E)-1 and its artificial (Z)-isomer
(Z)-1. Herein we describe a general method for the
stereoselective synthesis of both (E)-ΔAbu and (Z)-ΔAbu
amides and report the solid-phase total synthesis of bogorol A
[(E)-1] and its isomer (Z)-1. Intriguingly, preliminary
biological evaluation revealed that the geometrical isomer
(Z)-1 was equipotent to (E)-1.
We envisioned adopting the full solid-phase approach for the
total syntheses of (E)- and (Z)-1 in order to simplify the
operations and to facilitate a future comprehensive structure−
activity relationship study to search for optimized analogues
(Scheme 1).9,10 Resin-bound dodecapeptides (E)- and (Z)-2
were thus designed as the precursors of (E)- and (Z)-1,
respectively. In the synthetic direction, a single acidic treatment
would simultaneously realize release of (E)- or (Z)-1 from the
2-chlorotrityl resin and removal of all the acid labile protecting
groups (TBS of N-cap, Boc of D-Orn, D-Lys and L-Lys, and t-Bu
of D-Tyr) from the side chains. Protected peptides (E)- and
(Z)-2 would in turn be elongated by the combination of
standard Fmoc-based solid-phase peptide synthesis (SPPS)11
and on-resin construction of the (E)-ΔAbu and (Z)-ΔAbu
amides, respectively.
B
microbiome. Nosocomial infections caused by methicillin-
resistant Staphylococcus aureus (MRSA) and vancomycin-
resistant Enterococcus spp. (VRE), in particular, pose a global
threat to public health.1 New antibacterial lead structures that
show no cross-resistance with existing antibiotics are thus
urgently needed to guarantee future therapeutic efficacy.2
Bogorol A [(E)-1, Scheme 1] was isolated from cultures of
the marine bacterium Brevibacillus laterosporus collected in
Papua New Guinea and was reported to exhibit selective and
potent activity against MRSA [MIC (minimum inhibitory
concentration) = 2.5 μg/mL] and VRE (MIC = 9 μg/mL).3
One dehydroamino acid [(E)-ΔAbu] and four D-amino acids
(D-Orn, D-Lys, D-Leu, and D-Tyr) are incorporated within the
linear dodecapeptide sequence, which is further modified by
(2S,3S)-2-hydroxy-3-methylpentanoic acid (N-cap) at the N-
terminus and L-valinol (C-cap) at the C-terminus. These
nonproteinogenic structural units indicate the nonribosomal
origin of (E)-1.4 In addition, since the three amino groups of
the side chains contribute to the net positive charge of the
molecule, (E)-1 is classified as a cationic peptide antibiotic.
These peptide antibiotics are widespread in nature, are an
integral part of the innate immune systems,5 and have attracted
much attention as possible new-generation clinically useful
antibacterial agents due to their potent antibacterial activity,
broad antibiotic spectrum, and low propensity for rapid
emergence of resistance.6 Although many of these peptides
are known to kill bacteria by physically disrupting membrane
function, the precise mode of action of (E)-1 remains
unknown.
The well-known thermodynamic instability of an (E)-α,β-
dehydroamino acid residue in comparison to its (Z)-counter-
part makes its stereoselective construction highly challeng-
ing.12,13 In general, the 1,3-allylic strain of the (E)-
dehydroamino acid residue is higher than that of the (Z)-
The most unusual structural feature of (E)-1 is the presence
of thermodynamically unfavored (E)-2-amino-2-butenoic acid
[(E)-ΔAbu]. The conformation of the dehydroamino acid
residue is influenced by the β-substituent on a CαCβ double
Received: March 17, 2015
Published: April 13, 2015
© 2015 American Chemical Society
2170
DOI: 10.1021/acs.orglett.5b00769
Org. Lett. 2015, 17, 2170−2173

Organic Letters
Letter
a
Scheme 1. Structures and Retrosyntheses of Bogorol A and Its Isomer
a
TBS = tert-butyldimethylsilyl; Boc = tert-butoxycarbonyl.
derivative and increases when the substituted nitrogen (amide:
X = NHR2) is introduced at the C-terminus in place of the less
sterically bulky oxygen (ester: X = OR2) (Scheme 2a). To
develop a robust method for the stereoselective construction of
the (E)-ΔAbu amide, the synthesis of (E)-6 was pursued as a
model study (Scheme 2b).
dehydration proved to be problematic, an alternative approach
was sought for the synthesis of the model compound (E)-6.
In our total syntheses of nobilamides B and D,16 a new
method for the stereoselective construction of the thermody-
namically favored (Z)-ΔAbu ester was devised by employing
traceless Staudinger ligation.17,18 Thus, we applied this protocol
to the more challenging (E)-ΔAbu amide (E)-6 (Scheme 3).
Scheme 2. Attempted Synthesis of the (E)-ΔAbu Amide by
a
Scheme 3. Stereocontrolled Syntheses of the (E)- and (Z)-
ΔAbu Amides
Dehydration
a
a
DMSO = dimethyl sulfoxide; DMF = N,N-dimethylformamide; THF
= tetrahydrofuran, NMM = N-methylmorpholine; DME = 1,2-
dimethoxyethane.
a
EDC = 1-ethyl-3-(3-(dimethylamino)propyl)carbodiimide; PyBOP =
benzotriazol-1-yloxytripyrrolidinophosphonium hexafluorophosphate.
Before doing so, alkenyl azide (E)-14 was prepared by a four-
step sequence from ethyl but-2-ynoate 10. Specifically, regio-
and stereoselective hydrostannation19 of 10 and subsequent
iododestannation20 furnished alkenyl iodide (E)-11. Treatment
of iodide (E)-11 with NaN3, CuI, and ligand 12 in turn afforded
azide (E)-13, the ester moiety of which was hydrolyzed to
generate carboxylic acid (E)-14. Then, PyBOP21-promoted
condensation of (E)-14 with benzylamine stereospecifically
provided amide (E)-15. No isomerization would originate from
the absence of the N-terminal carbonyl group, which would
cyclize into the azlactone derivative (e.g., 8). Traceless
Staudinger ligation of the obtained azide (E)-15 occurred at
room temperature by simply mixing with phosphine 1616 in
aqueous THF for 24 h, furnishing the (E)-ΔAbu amide (E)-6
via ejection of (2-hydroxyphenyl)diphenylphosphine oxide
In accordance with Sai’s report,14 the EDC/CuCl2-promoted
syn-elimination of threonine methyl ester 3 selectively
generated (E)-ΔAbu ester (E)-5. However, the reaction of
threonine benzyl amide 4 under the same conditions was more
sluggish, slowly generating thermodynamically stable (Z)-6 as
the predominant product. Conversion of ester (E)-5 to amide
(E)-6 was also not successful. Saponification of (E)-5
proceeded without isomerization, but condensation between
carboxylic acid (E)-7 and benzylamine only afforded undesired
(Z)-6. In this reaction, the formation of the activated ester from
(E)-7 was presumably followed by cyclization into azlactone 8,
and equilibration between 8 and oxazole 9 resulted in
geometrical isomerization.15 Since seemingly straightforward
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Org. Lett. 2015, 17, 2170−2173

Organic Letters
Letter
a
Scheme 4. Total Syntheses of Bogorol A [(E)-1] and Its Isomer (Z)-1
a
DMAP = N,N-dimethyl-4-aminopyridine; Fmoc =9-fluorenylmethyloxycarbonyl; HBTU = O-(benzotriazol-1-yl)-N,N,N′,N′-tetramethyluronium
hexafluorophosphate; HOBt = 1-hydroxybenzotriazole; NMP = N-methylpyrrolidone; TFA = trifluoroacetic acid.
(78% yield). Moreover, the method was found to be applicable
for the synthesis of the (Z)-ΔAbu amide (Z)-6. Amidation of
acid (Z)-1416 and benzylamine using NMM and i-BuOCOCl,
and subsequent treatment of amide (Z)-15 with 16, led to
stereoselective generation of (Z)-6. The results shown in
Scheme 3 clarified the superiority of the traceless Staudinger
approach over conventional dehydration in terms of reliable
stereoselective outcomes and mild reaction conditions. Addi-
tionally, the order of stepwise amide bond formations, with the
C-terminus condensation preceding the Nα-Staudinger cou-
pling, permitted direct adoption of the method to the SPPS of
the targeted (E)- and (Z)-1.
Prior to the full solid-phase total syntheses of bogorol A
[(E)-1] and its isomer (Z)-1, N-cap 19, the reactive substrate
for the traceless Staudinger ligation, was prepared from the
known carboxylic acid 1722 and phosphinophenol 1823 using
EDC and DMAP (Scheme 4). Then, undecapeptide 21 was
elongated from the C-cap-bound chlorotrityl resin 20 by
employing Fmoc chemistry under microwave-assisted con-
ditions.24 Namely, 11 cycles of HBTU/HOBt25-mediated
peptide coupling and piperidine-promoted Nα-deprotection
using the component amino acid monomers [Fmoc-L-Leu-OH,
Fmoc-D-Tyr(t-Bu)-OH, Fmoc-L-Lys(Boc)-OH, Fmoc-D-Leu-
OH, Fmoc-L-Val-OH, Fmoc-D-Lys(Boc)-OH, Fmoc-L-Val-
OH, Fmoc-L-Val-OH, Fmoc-L-Ile-OH, Fmoc-D-Orn(Boc)-
OH, and Fmoc-L-Leu-OH] transformed 20 into 21. The
crucial installation of (E)-ΔAbu within the sequence was
achieved on-resin by the subsequent two steps. Acid (E)-14 was
attached to the N-terminus of 21 using PyBOP to produce
azide (E)-22, which was treated with N-cap 19 in aqueous THF
(0.029 M) for 3 days, delivering the fully protected bogorol A
[(E)-2]. Lastly, subjection of (E)-2 to 95% aqueous TFA
simultaneously removed the t-Bu, Boc, and TBS groups and
cleaved the peptide from the resin to release (E)-1 into
solution. After purification by the reversed-phase HPLC,
bogorol A [(E)-1] was obtained as the single isomer in 31%
total yield in 25 steps from 20. The non-natural (Z)-isomer of
bogorol A [(Z)-1] was constructed in the same manner by
using (Z)-14 instead of (E)-14 [20 → (Z)-2 → (Z)-1, 27%
total yield in 25 steps]. The exclusive formation of the (E)-
ΔAbu and (Z)-ΔAbu residues and excellent overall yields of
(E)- and (Z)-1 demonstrated the power and generality of the
traceless Staudinger approach for the assembly of complex
dehydro peptides. It is noteworthy that, in contrast to the facile
isomerization of the (E)-ΔAbu residue observed in Scheme 2,
the olefin geometry of (E)-1 was stable when incorporated into
the sequence: isomerization between (E)- and (Z)-1 was not
detected upon separate incubation of the two isomers in
aqueous buffer at 40 °C for 24 h.
1
Assignment of the H and 13C NMR peaks of (E)-1 was
problematic due to significant overlaps. Accordingly, the
authenticity of (E)-1 was confirmed after its derivatization
into the hexaacetylated compound (E)-23.3a The H and 13C
1
NMR spectra of synthetic (E)-23 were fully assigned and
shown to match with (E)-23 derived from the isolated natural
product 1.
A preliminary antibacterial study of (E)- and (Z)-1 was
carried out using methicillin-susceptible S. aureus (MSSA).
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Organic Letters
Letter
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remains to be elucidated, the equipotency of natural (E)-1 and
artificial (Z)-1 revealed the inconsequential nature of the olefin
geometry for exerting bioactivity.
In conclusion, we have achieved the total synthesis and
biological evaluation of antibiotic bogorol A [(E)-1] and its
isomer (Z)-1 for the first time. Traceless Staudinger ligation
was utilized for stereoselective construction of both the
thermodynamically unfavored (E)-ΔAbu amide of (E)-1 and
the favored (Z)-ΔAbu amide of (Z)-1. The developed method
was general and robust in comparison to the previous
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solid-phase route to the target compounds (E)- and (Z)-1
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ASSOCIATED CONTENT
■
(b) Bonauer, C.; Walenzyk, T.; Konig, B. Synthesis 2006, 1.
̈
(c) Kuranaga, T.; Sesoko, Y.; Inoue, M. Nat. Prod. Rep. 2014, 31, 514.
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S
* Supporting Information
Characterization data for all new compounds and experimental
procedures. This material is available free of charge via the
Internet at http://pubs.acs.org.
(16) Yamashita, T.; Matoba, H.; Kuranaga, T.; Inoue, M. Tetrahedron
2014, 70, 7746.
AUTHOR INFORMATION
■
(17) Representative works on the Staudinger ligation, see:
(a) Nilsson, B. L.; Kiessling, L. L.; Raines, R. T. Org. Lett. 2000, 2,
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Corresponding Author
*E-mail: inoue@mol.f.u-tokyo.ac.jp.
Notes
The authors declare no competing financial interest.
́ ́ ́
(f) Fernandez-Suarez, M.; Baruah, H.; Martinez-Hernandez, L.; Xie, K.
ACKNOWLEDGMENTS
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2012, 18, 14444. (h) Kosal, A. D.; Wilson, E. E.; Ashfeld, B. L. Angew.
Chem., Int. Ed. 2012, 51, 12036.
■
This research was financially supported by a Grant-in-Aid for
Scientific Research (A) to M.I. and a Grant-in-Aid for Young
Scientists (B) (JSPS) to T.K. We thank Prof. Kazuhisa
Sekimizu, Dr. Jyunichiro Yasukawa, and Dr. Hiroshi Hamamoto
(The University of Tokyo) for evaluation of antibacterial
activity.
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