Mycothiol synthesis by an anomerization reaction through endocyclic cleavage

Article abstract of DOI:10.3762/bjoc.12.35

Mycothiol is found in Gram-positive bacteria, where it helps in maintaining a reducing intracellular environment and it plays an important role in protecting the cell from toxic chemicals. The inhibition of the mycothiol biosynthesis is considered as a treatment for tuberculosis. Mycothiol contains an α-aminoglycoside, which is difficult to prepare stereoselectively by a conventional glycosylation reaction. In this study, mycothiol was synthesized by an anomerization reaction from an easily prepared β-aminoglycoside through endocyclic cleavage.

Full text of DOI:10.3762/bjoc.12.35

Mycothiol synthesis by an anomerization reaction through
endocyclic cleavage
Shino Manabe* and Yukishige Ito*
Letter
Open Access
Address:
Beilstein J. Org. Chem. 2016, 12, 328–333.
Synthetic Cellular Chemistry Lab, RIKEN, Hirosawa, Wako, Saitama
351-0198, Japan
doi:10.3762/bjoc.12.35
Received: 23 December 2015
Accepted: 11 February 2016
Published: 22 February 2016
Email:
Shino Manabe* - smanabe@riken.jp; Yukishige Ito* - yukito@riken.jp
* Corresponding author
Associate Editor: S. Flitsch
Keywords:
© 2016 Manabe and Ito; licensee Beilstein-Institut.
License and terms: see end of document.
anomerization; aminoglycoside; endocyclic cleavage reaction; inositol;
mycothiol
Abstract
Mycothiol is found in Gram-positive bacteria, where it helps in maintaining a reducing intracellular environment and it plays an im-
portant role in protecting the cell from toxic chemicals. The inhibition of the mycothiol biosynthesis is considered as a treatment for
tuberculosis. Mycothiol contains an α-aminoglycoside, which is difficult to prepare stereoselectively by a conventional glycosyla-
tion reaction. In this study, mycothiol was synthesized by an anomerization reaction from an easily prepared β-aminoglycoside
through endocyclic cleavage.
Introduction
Tuberculosis is an infectious disease and has had a high death NMR analyses and computational calculations [11,12].
rate over the past few decades [1-4]. The occurrence of Recently, N-acyl variants of MSH homologs, such as formyl,
multiple-drug-resistant (MDR), extensive-drug-resistant (EDR), propanoyl, and succinoyl, have been reported [13-15].
and totally drug-resistant (TDR) pathogens has increased the
need for new drug candidates for treating tuberculosis.
Mycothiol (MSH) 1 is the main low-molecular-weight thiol
found in most actinomycetes, including Mycobacteria and
Streptomycetes [5-10]. It consists of an N-acetylcysteine, a
D-glucosamine, and a D-myo-inositol moiety (Figure 1).
D-Glucosamine is α-linked to D-myo-inositol at the 1-position,
and N-acetylcysteine is linked to the amino group of D-glucos-
Figure 1: Structure of mycothiol 1.
amine. The conformation of MSH has been investigated by
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Beilstein J. Org. Chem. 2016, 12, 328–333.
Gram-negative bacteria and most Eukaryotes utilize glutathione Due to the limited availability of MSH from M. smegmatis cell
as a low-molecular-weight thiol for maintaining a reducing culture (<1.5 mg of MSH from 1 L culture) [20], the chemical
environment in the cytosol. Gram-positive bacteria including synthesis of MSH is highly desired. Bewley et al. and Lee and
actinomycetes lack glutathione, instead, MSH is found as the Rosazza independently reported the synthesis of MSH and de-
major low-molecular-weight thiol. It is considered that MSH is termined the absolute stereochemistry of the glucosamine and
required for maintaining a reducing intracellular environment in cysteine moieties [21,22]. Hung also reported the utility of the
Gram-positive bacteria, similar to glutathione in eukaryotes and 2-azido-2-deoxyglycosyl donor and resolved inositol isomers in
Gram-negative bacteria. MSH undergoes metal-catalyzed a recent MSH synthesis [23]. Knapp et al. reported an intramo-
autoxidation more rapidly than glutathione [16]. The biosyn- lecular aglycone delivery method in order to achieve complete
thetic pathway of MSH has been well investigated; MSH is syn- α-stereoselectivity [24]. The Ni(4-F-PhCN)4(OTf)2-catalyzed
thesized from 1-inositol phosphate and uridine diphosphate synthesis of N-substituted benzylideneaminoglycosides has also
N-acetylglucosamine (UDP-GlcNAc) in five steps [15]. It is been achieved to fabricate MSH [25]. The α-stereoselective for-
used by mycobacteria for protection against foreign electrophil- mation of aminoglycoside is the crucial step in MSH synthesis.
ic agents such as oxidants, radicals, and drugs. In the detoxifica- However, except for the intramolecular aglycone delivery
tion pathway, MSH reacts with alkylating reagents and the re- method developed by Knapp et al., complete α-stereoselective
sulting S-conjugates are subsequently cleaved at the amide bond glycosylation reactions were difficult in mycothiol synthesis.
by MSH S-conjugate amidase (Scheme 1) [4-10]. After The complete α-stereoselective glycosylation reaction of amino-
cleavage, the N-acetylcysteine S-conjugate is transported out of glycosides is still generally difficult at this moment [26-28].
the cell, while the inositol–glucosamine conjugate is recycled to
afford MSH. MSH also plays an important role in the growth Oscarson and our group recently demonstrated that reactions of
and survival of Mycobacterium tuberculosis. Because MSH-de- pyranosides with N-acetyl 2,3-trans-carbamate groups exhib-
pendent pathways are not found in eukaryotes, the enzymes ited complete anomerization from β-glycoside to α-glycoside in
involved may be considered as novel antimicrobacterial targets, the presence of a weak Lewis acid through an endocyclic
especially for tuberculosis, and several compounds with cleavage reaction [29-33]. We showed evidence of the endo-
inhibitory activity have been synthesized [17,18]. In addition, cyclic cleavage reaction by trapping linear cations through
another function of MSH was recently reported; it is involved reduction, and intramolecular Friedel–Crafts reaction [29,30]
in the biosynthesis of lincomycin A, a sulfur-containing (Scheme 2). In particular, the reaction of pyranosides bearing
lincosamide antibiotic [19].
acetyl substituents on the carbamate groups showed complete
Scheme 1: Detoxification pathway mediated by MSH.
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Beilstein J. Org. Chem. 2016, 12, 328–333.
Scheme 2: Anomerization via endocyclic cleavage.
anomerization [32]. We expected that anomerization via endo- tion, was converted to α-glycoside 4, by introducing an N-acetyl
cyclic cleavage would be useful for mycothiol synthesis.
2,3-trans-carbamate group (Scheme 3) and by conducting an
anomerization reaction.
Results and Discussion
Based on the results of our previous study, we expected that an The glycosylation reaction of phthalimido-protected glucos-
anomerization would be useful for the stereoselective synthesis amine thioglycoside 5 with inositol 6 [24] afforded β-linked
of α-aminoglycosides, which is normally difficult by conven- pseudo-disaccharide 2 in 90% yield (Scheme 4). After
tional glycosylation reactions. β-Glycoside 2, which is synthe- removing the phthaloyl and acetyl groups by using ethylenedi-
sized by assistance from the phthalimide group in the 2-posi- amine in dimethylformamide (DMF), a carbamate group was
Scheme 3: Outline of mycothiol synthesis by anomerization.
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Beilstein J. Org. Chem. 2016, 12, 328–333.
Scheme 4: Synthesis of a pseudodisaccharide by an anomerization reaction.
introduced using triphosgene in the presence of NaHCO3. H2/Pd(OH)2/C in AcOH/dioxane/H2O. Although it was re-
Acetylation of both the hydroxy and carbamate groups was ported that the cysteine moiety was introduced by 1-[bis(di-
carried out using acetic anhydride in pyridine in the presence of methylamino)methylene]-1H-1,2,3-triazolo[4,5-b]pyridinium
4-dimethylaminopyridine (DMAP). Finally, β-glycoside 3 was 3-oxid hexafluorophosphate (HATU) [23,24], we found that
completely anomerized to the corresponding α-anomer 4 purification of the product was rather difficult, especially for
in quantitative yield in the presence of two equivalents of >50 mg-scale reactions. The reason for the low yield is proba-
BF3·OEt2 in CH3CN [33] within 30 min at −30 °C.
bly the low solubility of the product in the reported solvent
system during column chromatography (CHCl3/MeOH/AcOH),
With the α-linked inositol 4 in hand, the synthesis of MSH was and azabenzotriazole from HATU was difficult to remove.
completed as follows (Scheme 5): the carbamate and acetyl Instead, ethyl (hydroxyimino)cyanoacetate (COMU) [34] cou-
groups were removed by alkaline hydrolysis to give known pling gave product 9 in 80% yield after reversed-phase column
compound 8 [24]; then, benzyl groups were removed by chromatography purification (H2O/MeOH). Finally, Boc
Scheme 5: Mycothiol synthesis from pseudo-disaccharide 4.
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Conclusion
We demonstrated a MSH synthesis using an anomerization
reaction through endocyclic cleavage in the presence of a weak
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S. M. received support through a Grant-in-Aid for Scientific
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Promotion of Science, Yamada Science Foundation, Takeda
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