Concise synthesis of an arabinofuranose hexasaccharide present in the cell wall of Mycobacterium tuberculosis

Mycobacterium cell wall consists of three major polysaccharide portions and arabinofuranose (Araf) is present in two of the major portions, arabinogalactan (AG) and lipoarabinomannan (LAM). A peculiar Araf hexasaccharide possessing two β-linked Araf units are present in both AG and LAM polysaccharides. Herein, we report an efficient and concise synthesis of this Araf hexasaccharide using single starting 3,5-TIPDS protected Araf thioglycoside precursor. Double β-glycosylation was achieved using strained cyclic 2-p-methoxybenzyl-3,5-TIPDS Araf thioglycoside donor.     

Synthesis of galactofuranose-containing acceptor substrates for mycobacterial galactofuranosyltransferases

The major structural component of the cell wall in Mycobacterium tuberculosis, infection by which causes tuberculosis, is the mycolyl-arabinogalactan (mAG) complex. This large glycoconjugates has at its core a backbone of approximately 30 D-galactofuranose (Gal(f)) residues that are linked to peptidoglycan by way of a linker disaccharide containing L-rhamnose and 2-acetamido-2-deoxy-D-glucose. Recent studies have supported a model of galactan biosynthesis in which the entire structure is assembled by the action of two bifunctional galactofuranosyltransferases. These biochemical investigations were made possible, in part, by access to a panel of oligosaccharide fragments of the mAG complex (1-12), the synthesis of which we describe here. An early key finding in this study was that the iodine-promoted cyclization of galactose diethyl dithioacetal (19) in the presence of an alcohol solvent led to the formation Gal(f) glycosides contaminated with no pyranoside isomer, thus allowing the efficient preparation of furanoside derivatives of this monosaccharide. The synthesis of disaccharide targets 1, 2, 11 and 12 proceeded without difficulty through the use of thioglycoside donors and octyl glycoside acceptors, both carrying benzoyl protection. In the synthesis of the tri- and tetrasaccharides 3-6, we explored routes in which the molecule was assembled from the reducing to nonreducing end, and the reverse. The latter approach was found to be preferable for the preparation of 6, and in the case of 3 and 4, this strategy allowed the development of efficient one-pot methods for their synthesis. We have also carried out the first synthesis of three mAG fragments (8-10) consisting of the linker disaccharide further elaborated with one, two or three Gal(f) residues. A key step in the synthesis of these target compounds was the coupling of a protected linker disaccharide derivative (58) with a mono-, di-, or trigalactofuranosyl thioglycoside (17, 54, or 53, respectively).     

L-薄荷基-β-D-半乳糖苷的合成新方法

首次报道了硫代糖苷法合成L-薄荷基-β-D-半乳糖苷。半乳糖经过乙酰化、硫代、脱乙酰基和苯甲酰基化反应,转变成供体异丙基-2,3,4,6-四-O-苯甲酰基-1-硫-β-D-半乳糖苷。该供体与L-薄荷醇偶联及脱保护,获得了L-薄荷基-β-D-半乳糖苷。所有的产品通过NMR、MS等方法进行了结构表征。通过TG方法,探讨了所合成的L-薄荷基-β-D-半乳糖苷的热失重过程。     

Characterization of a distinct arabinofuranosyltransferase in Mycobacterium smegmatis

The D-arabinans in Mycobacterium are essential, extraordinarily complex entity comprised of d-arabinofuranose residues which are rarely found in nature. Despite the well-recognized importance of the mycobacterial arabinan, delineation of the arabinosylation process has been severely hampered due to lack of positively identified arabinosyltransferases. Identification of genes involved in arabinan biosynthesis entailed the use of ethambutol (EMB), a first-line antituberculosis agent that is known to inhibit cell wall arabinan synthesis. The three genes (embA, embB, and embC) encode novel membrane proteins, implicated as the only known mycobacterial arabinosyltransferases to this date. We have now adapted a multifaceted approach involving development of convenient arabinosyltransferase assay using novel synthetic acceptors to identify arabinosyltransferase/s that will be distinct from the Emb proteins. In our present work, Mycobacterium smegmatis mc(2) 155 (WTMsm) was used as a model to study the biosynthesis of cell wall arabinan. In an in vitro assay, we demonstrate that transfer of only alpha-Araf had occurred from decaprenylphosphoryl-D-arabinofuranose (DPA) on a newly synthesized branched acceptor [alpha-D-Araf](2)-3,5-alpha-D-Araf-(1-->5)-alpha-d-Araf-(1-->5)-alpha-D-Araf with an octyl aglycon. Higher molecular weight (up to Ara(10)) oligomers were also detected in a parallel reaction using cold phosphoribosepyrophosphate (pRpp). Matrix-assisted laser desorption ionization time-of-flight tandem mass spectrometry (MALDI-TOF MS/MS) analysis of these products revealed that isomeric products were formed and initiation and elongation of arabinan can occur either on the 5-arm or 3-arm of the branched 3,5-alpha-D-Araf. Individual embA, embB, and embC knockout strains retained this alpha-1,5 arabinosyltransferase activity, and the activity was partially inhibited by ethambutol. This particular enzyme function is distinct from the function of the Emb proteins.     

Synthesis of the C(18)-C(34) fragment of amphidinolide C and the C(18)-C(29) fragment of amphidinolide F

A convergent synthesis of the C(18)-C(34) fragment of amphidinolide C and the C(18)-C(29) fragment of amphidinolide F is reported. The approach involves the synthesis of the common intermediate tetrahydrofuranyl-β-ketophosphonate via cross metathesis, Pd(0)-catalyzed cyclization, and hydroboration-oxidation. The β-ketophosphonate was coupled to three side chain aldehydes using a Horner-Wadsworth-Emmons (HWE) olefination reaction to give dienones, which were reduced with l-selectride to give the fragments of amphidinolide C and F.     

Lewis acids as α-directing additives in glycosylations by using 2,3-O-carbonate-protected glucose and galactose thioglycoside donors based on preactivation protocol

Catalytic or stoichiometric amounts of Lewis acids were found to be very effective α-directing additives in the stereoselective glycosylations of diverse 2,3-O-carbonate-protected glucose and galactose thioglycoside donors by preactivation protocol. The poor stereoselectivities of 4,6-di-O-acetyl-2,3-O-carbonate protected thioglycoside donors in glycosyl coupling reactions were greatly improved, and excellent α-stereoselectivities were achieved by the addition of 0.2 equiv of BF(3)·OEt(2). On the other hand, the β-selectivities of 4,6-di-O-benzyl-2,3-O-carbonate-protected thioglucoside donor toward glycosylations were reversed completely to the α-selectivities by the use of 1 equiv of SnCl(4), making the stereoselectivity controllable. Furthermore, the poor stereoselectivities of 4,6-di-O-benzyl-2,3-O-carbonate-protected thiogalactoside donor in glycosylations were also improved by using SnCl(4) as additive.     

Convergent synthesis of the tetrasaccharide repeating unit related to the O-antigenic polysaccharide of Escherichia coli 78

A convergent synthesis of the tetrasaccharide repeating unit of the O-antigenic cell wall polysaccharide of Escherichia coli 78, as the corresponding methyl glycoside (I), is being reported. It involved stereoselective glycosidation of a β-linked mannodisaccharide acceptor with a β-linked glucosamine based disaccharide thioglycoside donor, which were prepared from the corresponding functionalised monosaccharide based glycosyl donors and acceptors. The resulting tetrasaccharide derivative was finally converted to (I) by selective deprotection and also by global protection and deprotection techniques.     

Synthesis of glycosaminoglycan oligosaccharides. Part 5: Synthesis of a putative heparan sulfate tetrasaccharide antigen involved in prion diseases

The synthesis of the putative prion-associated heparan sulfate tetrasaccharide containing two d-glucuronic acid units is reported. Key to the synthesis were the differentiation of the N-acetylated and N-unsubstituted glucosamine units, the high-yielding glucosylation at O-4 of an N-acetyl-d-glucosamine derivative and the α-selective glycosylation of the 4′-OH group of a β-d-GlcpA-(1→4)-d-GlcpNAc disaccharide building block with a disaccharide thioglycoside donor.     

Synthesis of putative chain terminators of mycobacterial arabinan biosynthesis

The synthesis of a variety of arabinose derivatives that have been modified at C-5 was achieved from d-arabinose. The 5-fluoro and 5-methoxy compounds were converted into the corresponding farnesyl phosphodiesters as putative chain terminators of mycobacterial arabinan biosynthesis. Biological testing of these materials revealed no effective anti-mycobacterial activity.     

Synthesis of a natural oligosaccharide antibiotic active against Helicobacter pylori

An oligosaccharide active against Helicobacter pylori was synthesized in a highly efficient manner for the first time. The anti-H. pylori oligosaccharide structure is a core-2 branched-type oligosaccharide with a characteristic alpha-N-acetylglucosamine at the nonreducing end. The oligosaccharide was synthesized from the nonreducing end to the reducing end, with an N-benzyl-2,3-oxazolidinone-carrying glycosyl donor used to introduce an alpha-N-acetylglucosamine at the nonreducing end. Complete chemoselective activation of a bromo sugar in the presence of a thioglycoside acceptor was achieved, and the use of 2,6-dimethylphenyl thioglycoside prevented the aglycon transfer observed when the corresponding phenyl thioglycoside is used as an acceptor.     

Synthesis of a Heptasaccharide,Structurally Related to the Phytoelicitor Active Glucan of Phytophthora Megasperma F.SP. Glycinea

Abstract

A synthesis is described of the heptasaccharide 1, which may form part of the phytoelicitor-active glucan of Phytopkthora megasperma f. sp. glycinea. Silver triflate was used as the promoter in Koenigs-Knorr type condensations using glycosyl bromides, each with a participating benzoyl group in the 2-position, for the synthesis of the smaller oligosaccharide fragments. For joining the larger ones, methyl triflate was used as the promoter and an oligosaccharide thioglycoside carrying a participating benzoyl group in the 2-position was used as the glycosyl donor.     

A novel O-fucosylation strategy preactivated by (p-Tol)2SO/Tf2O and its application for the synthesis of Lewis blood group antigen Lewisa

Based on a preactivation strategy using (p-Tol)₂SO/Tf₂O, a new O-fucosylation method with thioglycoside as donor under mild conditions was reported. High yields and excellent α-stereoselectivities of the fucosylation were obtained with secondary sugar alcohol as acceptors. Moreover, the novel fucosylation strategy was successfully applied to the synthesis of an important Lewis blood group antigen Lewis^a, which was obtained in nine linear steps with 27% overall yield.     

First synthesis of the beta-D-rhamnosylated trisaccharide repeating unit of the O-antigen from Xanthomonas campestris pv. campestris 8004

The trisaccharide repeating unit of the O-antigen of the lipopolysaccharide from Xanthomonas campestris pv. campestris 8004, a pathogen of cruciferous crops, presents some structural features that renders it a challenging synthetic target: the presence of a beta-D-rhamnosidic linkage, the steric crowd on a 1,2-cis-diglycosylated D-rhamnose, and finally the noncommercial availability of its monosaccharide constituents. The synthesis of this trisaccharide as methyl glycoside has been accomplished by exploiting a strategy whose key steps were the sequential beta-D-rhamnosylation with a 2-O-benzylsulfonyl-N-phenyltrifluoroacetimidate donor, debenzylsulfonylation, and coupling with a D-Fucp3NAc thioglycoside donor.     

Synthesis and Biological Evaluation of the Forssman Antigen Pentasaccharide and Derivatives by a One‐Pot Glycosylation Procedure

The synthesis and biological evaluation of the Forssman antigen pentasaccharide and derivatives thereof by using a one‐pot glycosylation and polymer‐assisted deprotection is described. The Forssman antigen pentasaccharide, composed of GalNAcα(1,3)GalNAcβ(1,3)Galα(1,4)Galβ(1,4)Glc, was recently identified as a ligand of the lectin SLL‐2 isolated from an octocoral Sinularia lochmodes. The chemo‐ and α‐selective glycosylation of a thiogalactoside with a hemiacetal donor by using a mixture of Tf₂O, TTBP and Ph₂SO, followed by activation of the remaining thioglycoside, provided the trisaccharide at the reducing end in a one‐pot procedure. The pentasaccharide was prepared by the α‐selective glycosylation of the N‐Troc‐protected (Troc=2,2,2‐trichloroethoxycarbonyl) thioglycoside with a 2‐azide‐1‐hydroxyl glycosyl donor, followed by glycosidation of the resulting disaccharide at the C3 hydroxyl group of the trisaccharide acceptor in a one‐pot process. We next applied the one‐pot glycosylation method to the synthesis of pentasaccharides in which the galactosamine units were partially and fully replaced by galactose units. Among the three possible pentasaccharides, Galα(1,3)GalNAc and Galα(1,3)Gal derivatives were successfully prepared by the established method. An assay of the binding of the synthetic oligosaccharides to a fluorescent‐labeled SLL‐2 revealed that the NHAc substituents and the length of the oligosaccharide chain were both important for the binding of the oligosaccharide to SLL‐2. The inhibition effect of the oligosaccharide relative to the morphological changes of Symbiodinium by SLL‐2, was comparable to their binding affinity to SLL‐2. In addition, we fortuitously found that the synthetic Forssman antigen pentasaccharide directly promotes a morphological change in Symbiodinium. These results strongly indicate that the Forssman antigen also functions as a chemical mediator of Symbiodinium.     

Explorations on the total synthesis of the unusual marine alkaloid chartelline A

In work directed toward a total synthesis of chartelline A (1a), a strategy was investigated to construct the 10-membered ring of this marine alkaloid via an intramolecular aldehyde/beta-lactam cyclocondensation to form the macrocyclic enamide functionality. Therefore, spiro-beta-lactam and imidazole fragments were first prepared. Tribromooxindole beta-lactam 24 was synthesized from commercially available 5-nitroisatin (18) in seven steps and 30% overall yield via a Staudinger ketene-imine [2 + 2]-cycloaddition strategy. The requisite 2-bromoimidazole subunit 40 bearing a terminal alkyne and a masked aldehyde was efficiently prepared from the readily available imidazole ester 25 in 10 steps. With both advanced intermediates available, the addition of the lithium acetylide generated from 2-bromoimidazole subunit 40 to the gamma-lactam carbonyl group of N-Boc-tribromooxindole 24 was investigated, affording the desired N-Boc-aminal 41. Hydrolysis of the acetonide moiety of 41, followed by oxidative cleavage of the resulting diol, gave the aldehyde 42. Unfortunately, treatment of aldehyde 42 with p-toluenesulfonic acid did not give the desired 10-membered macrocyclic (Z)-enamide 46, but rather the highly unsaturated seven-membered ring compound 44.     

Stereoselective synthesis of a fragment of mycobacterial arabinan

Strategies for the stereoselective synthesis of mycobacterial arabinan were explored. Arabinofuranosyl donors with various protective groups were screened in terms of suitability for beta-(1,2-cis)-selective glycosylation. The protective group was found to affect the stereoselectivity of arabinofuranosylation. Beta-selectivity was drastically enhanced by using donors protected with 3,5-TIDPS, possibly due to conformational constraints on the furanose ring. Synthesis of heptaarabinofuranoside was then performed to demonstrate the practicality of this methodology. [reaction: see text].     

β-Kdo乙硫苷供体的合成

以草酰乙酸和D-阿拉伯糖为原料,经缩合-脱羧,乙酰化,甲酯化和乙硫苷化等4步反应合成了β-Kdo乙硫苷供体,总收率24%,立体选择性单一,其结构经~1H NMR和~(13)C NMR确证。     

Sequence analysis of alginate-derived oligosaccharides by negative-ion electrospray tandem mass spectrometry

Negative-ion electrospray tandem mass spectrometry (ES-MS/MS) with collision-induced dissociation (CID) is attempted for sequence determination of alginate oligosaccharides, derived from polyanionic alginic acid, polymannuronate, and polyguluronate by partial depolymerization using either alginate lyase or mild acid hydrolysis. Sixteen homo- and hetero-oligomeric fragments were obtained after fractionation by gel-filtration and strong anion exchange high performance liquid chromatography. The product-ion spectra of these alginate oligosaccharides were dominated by intense B-, C-, Y-, and Z-type ions together with (0,2)A- and (2,5)A-ions of lower intensities. Internal mannuronate residues (M) produce weak but specific decarboxylated Z(int)-ions (Z(int) - 44 Da; int: denotes internal), which can be used for distinction of M and a guluronate residue (G) at an internal position. A reducing terminal M or G, although neither gives rise to a specific ion, can be identified by differences in the intensity ratio of fragment ions of the reducing terminal residue [(2,5)A(red)]/[(0,4)A(red)] (red: denotes reducing terminal).     

Synthesis of the Trisaccharide Repeating Unit of the K-Antigen from Klebsiella Type-63

Abstract

The benzyl substituted ethyl thioglycoside of L-fucose was found to be a more reactive donor compared to 2-O-benzyl substituted p-tolyl thioglycoside of D-galactose. Using the benzyl substituted ethyl thioglycoside of L-fucose (8), as a donor and the suitably substituted p-tolyl thioglycoside of D-galactose (7) as acceptor, the p-tolyl thioglycoside of the disaccharide, 9, was prepared. This disaccharide donor was allowed to react with a suitably protected galactopyranosyluronic acid acceptor, 16, to give the trisaccharide repeating unit of the K-antigen from Klebsiella type 63.     

Synthesis of a tetrasaccharide corresponding to the teichoic acid from the cell wall of Streptomyces sp. VKM Ac-2275

A concise chemical synthesis of a tetrasaccharide found in the teichoic acid from the cell wall of Streptomyces sp. VKM Ac-2275 was achieved in high yield. A [2+2] block synthetic strategy has been adopted for the construction of the target tetrasaccharide by exploiting the orthogonal property of a thioglycoside. For the first time, the 2-(4-methoxyphenoxy) ethyl group has been used as the anomeric protecting group to make the glycone moiety with a readily available linker for its conjugation to a protein without destroying the cyclic structure at the reducing end. Yields were high in all of the intermediate steps.