Dendritic cell‐specific intercellular adhesion molecule‐3‐grabbing nonintegrin (DC‐SIGN) and Langerin are C‐type lectins of dendritic cells (DCs) that share a specificity for mannose and are involved in pathogen recognition. HIV is known to use DC‐SIGN on DCs to facilitate transinfection of T‐cells. Langerin, on the contrary, contributes to virus elimination; therefore, the inhibition of this latter receptor is undesired. Glycomimetic molecules targeting DC‐SIGN have been reported as promising agents for the inhibition of viral infections and for the modulation of immune responses mediated by DC‐SIGN. We show here for the first time that glycomimetics based on a mannose anchor can be tuned to selectively inhibit DC‐SIGN over Langerin. Based on structural and binding studies of a mannobioside mimic previously described by us ( 2 ), a focused library of derivatives was designed. The optimized synthesis gave fast and efficient access to a group of bis(amides), decorated with an azide‐terminated tether allowing further conjugation. SPR inhibition tests showed improvements over the parent pseudomannobioside by a factor of 3–4. A dimeric, macrocyclic structure ( 11 ) was also serendipitously obtained, which afforded a 30‐fold gain over the starting compound ( 2 ). The same ligands were tested against Langerin and found to exhibit high selectivity towards DC‐SIGN. Structural studies using saturation transfer difference NMR spectroscopy (STD‐NMR) were performed to analyze the binding mode of one representative library member with DC‐SIGN. Despite the overlap of some signals, it was established that the new ligand interacts with the protein in the same fashion as the parent pseudodisaccharide. The two aromatic amide moieties showed relatively high saturation in the STD spectrum, which suggests that the improved potency of the bis(amides) over the parent dimethyl ester can be attributed to lipophilic interactions between the aromatic groups of the ligand and the binding site of DC‐SIGN. 相似文献
A highly efficient cap‐exchange approach for preparing compact, dense polyvalent mannose‐capped quantum dots (QDs) has been developed. The resulting QDs have been successfully used to probe multivalent interactions of HIV/Ebola receptors DC‐SIGN and DC‐SIGNR (collectively termed as DC‐SIGN/R) using a sensitive, ratiometric Förster resonance energy transfer (FRET) assay. The QD probes specifically bind DC‐SIGN, but not its closely related receptor DC‐SIGNR, which is further confirmed by its specific blocking of DC‐SIGN engagement with the Ebola virus glycoprotein. Tuning the QD surface mannose valency reveals that DC‐SIGN binds more efficiently to densely packed mannosides. A FRET‐based thermodynamic study reveals that the binding is enthalpy‐driven. This work establishes QD FRET as a rapid, sensitive technique for probing structure and thermodynamics of multivalent protein–ligand interactions. 相似文献
A highly efficient cap‐exchange approach for preparing compact, dense polyvalent mannose‐capped quantum dots (QDs) has been developed. The resulting QDs have been successfully used to probe multivalent interactions of HIV/Ebola receptors DC‐SIGN and DC‐SIGNR (collectively termed as DC‐SIGN/R) using a sensitive, ratiometric Förster resonance energy transfer (FRET) assay. The QD probes specifically bind DC‐SIGN, but not its closely related receptor DC‐SIGNR, which is further confirmed by its specific blocking of DC‐SIGN engagement with the Ebola virus glycoprotein. Tuning the QD surface mannose valency reveals that DC‐SIGN binds more efficiently to densely packed mannosides. A FRET‐based thermodynamic study reveals that the binding is enthalpy‐driven. This work establishes QD FRET as a rapid, sensitive technique for probing structure and thermodynamics of multivalent protein–ligand interactions. 相似文献
Multivalent binding is a key for many critical biological processes and unique recognition and specificity in binding enables many of different glycans and proteins to work in a great harmony within the human body. In this study, the binding kinetics of synthetic glycopolypeptides to the dendritic cell lectin DC‐SIGN and their inhibition potential for DC‐SIGN interactions with the gp120 envelope glycoprotein of HIV‐1 (gp120) are investigated.
Eight carbohydrate‐modified perylene bisimides ( PBI‐4 lac‐2 lac , PBI‐4 lac‐2 Man , PBI‐4 lac‐2 Gal , PBI‐4 lac‐2 Mal , PBI‐4 Man‐2 Man , PBI‐4 Man‐2 lac , PBI‐4 Man‐2 Gal and PBI‐4 Man‐2 Mal ) were synthesized, and the following predetermined supramolecular chirality rule was found: perylene bisimides modified with disaccharides (D ‐lactose and D ‐maltose) at the imide position generated right‐handed chirality, and those modified with monosaccharides (D ‐mannose and D ‐galactose) generated left‐handed chirality, when D ‐lactose or D ‐mannose was substituted in the bay positions of perylene bisimides with amide bonds as the linking spacers. These results may be because of the difference in the stacking angle of the perylene bisimide backbones induced by the steric effect and the additional hydrogen bonds between the disaccharide residues. This study provides an important design rule for predetermined chiral self‐assembly of perylene bisimides. 相似文献
Glycoside hydrolase family 99 (GH99) was created to categorize sequence‐related glycosidases possessing endo‐α‐mannosidase activity: the cleavage of mannosidic linkages within eukaryotic N‐glycan precursors (Glc1–3Man9GlcNAc2), releasing mono‐, di‐ and triglucosylated‐mannose (Glc1–3‐1,3‐Man). GH99 family members have recently been implicated in the ability of Bacteroides spp., present within the gut microbiota, to metabolize fungal cell wall α‐mannans, releasing α‐1,3‐mannobiose by hydrolysing αMan‐1,3‐αMan→1,2‐αMan‐1,2‐αMan sequences within branches off the main α‐1,6‐mannan backbone. We report the development of a series of substrates and inhibitors, which we use to kinetically and structurally characterise this novel endo‐α‐1,2‐mannanase activity of bacterial GH99 enzymes from Bacteroides thetaiotaomicron and xylanisolvens. These data reveal an approximate 5 kJ mol?1 preference for mannose‐configured substrates in the ?2 subsite (relative to glucose), which inspired the development of a new inhibitor, α‐mannopyranosyl‐1,3‐isofagomine (ManIFG), the most potent (bacterial) GH99 inhibitor reported to date. X‐ray structures of ManIFG or a substrate in complex with wild‐type or inactive mutants, respectively, of B. xylanisolvens GH99 reveal the structural basis for binding to D ‐mannose‐ rather than D ‐glucose‐configured substrates. 相似文献
The D ‐manno‐tetrahydroimidazopyridine‐2‐phosphonate 11 was prepared via a high‐yielding Pd(PPh3)4‐catalysed diphenylphosphonylation of the manno‐iodoimidazole 12 , followed by transesterification to the diethyl phosphonate 14 and dealkylation, providing 11 in eight steps from the thionolactam 1 and in an overall yield of 15%. Alternatively, a more highly convergent synthesis based on the HgCl2/Et3N‐promoted condensation of the thionolactam 1 with the α‐aminophosphonate 24 in THF led to 11 in four steps and in the same overall yield. In the presence of HgCl2/Et3N, the thionolactam 1 reacted at 80° with 2‐methoxyethanol to provide 66% of a 64 : 36 mixture of the gluco‐ and manno‐iminoethers 29 / 30 . Performing the reaction at 22° yielded preferentially the gluco‐isomer 29 (86%, 84 : 16). 相似文献
The D ‐gluco‐isoquinuclidines 3 and 4 were prepared and tested as inhibitors of the β‐glucosidases from Caldocellum saccharolyticum and from sweet almonds; the results are compared to the inhibition of snail β‐mannosidase by the D ‐manno‐isoquinuclidines 1 and 2 . Exploratory experiments in the racemic series showed that treatment of the ester epoxide 6 with benzyl alcoholates leads only to epimerisation, transesterification, and formation of the cyclopropane 9 . Ring opening of the reduced epoxide 13 by NaN3 proceeded regioselectively to provide 14 . Treatment of the C(6)? O‐triflate 16 with AcOCs induced a rearrangement; the reaction with NaN3 gave the C(5)‐azido derivative 14 . The acetoxy triflate 18 , however, reacted with AcOCs to provide the desired gluco‐isoquinuclidine 19 . Similarly, the enantiomerically pure acetoxy triflate 22 provided the D ‐gluco‐isoquinuclidine 24 , which was reduced and deprotected to provide 3 and 4 . The deoxy analogues 30 and 31 were obtained by reductive deiodination of the iodide 27 , derived from 22 . The D ‐gluco‐isoquinuclidines 3, 4, 30 , and 31 are much weaker inhibitors of β‐glucosidases than the D ‐manno‐analogues 1 and 2 of snail β‐mannosidase. The N‐benzyl derivative 3 is a weaker inhibitor than the N‐unsubstituted analogue in the gluco‐series, while it is a much stronger inhibitor in the manno‐series. A consideration of the pKHA values of the isoquinuclidines 1 – 4 and the pH value of the enzyme assays suggests that the D ‐gluco‐isoquinuclidines are poor mimics of the shape of a reactive, enzyme‐bound gluco‐conformer, while the D ‐manno‐analogues are reasonably good mimics of a reactive, enzyme‐bound manno‐conformer. The inhibition results may also suggest that the glycosidase induced lengthening of the scissile bond and rehybridisation of the anomeric centre are more strongly correlated with the change of the ground‐state conformation during hydrolysis of β‐D ‐glucopyranosides than of β‐D ‐mannopyranosides. 相似文献
Racemic and enantiomerically pure manno‐configured isoquinuclidines were synthesized and tested as glycosidase inhibitors. The racemic key isoquinuclidine intermediate was prepared in high yield by a cycloaddition (tandem Michael addition/aldolisation) of the 3‐hydroxy‐1‐tosyl‐pyridone 10 to methyl acrylate, and transformed to the racemic N‐benzyl manno‐isoquinuclidine 2 and the N‐unsubstituted manno‐isoquinuclidine 3 (twelve steps; ca. 11% from 10 ). Catalysis by quinine of the analogous cycloaddition of 10 to (?)‐8‐phenylmenthyl acrylate provided a single diastereoisomer in high yield, which was transformed to the desired enantiomerically pure D ‐manno‐isoquinuclidines (+)‐ 2 and (+)‐ 3 (twelve steps; 23% from 10 ). The enantiomers (?)‐ 2 and (?)‐ 3 were prepared by using a quinidine‐promoted cycloaddition of 10 to the enantiomeric (+)‐8‐phenylmenthyl acrylate. The N‐benzyl D ‐manno‐isoquinuclidine (+)‐ 2 is a selective and slow inhibitor of snail β‐mannosidase. Its inhibition strength and type depends on the pH (at pH 4.5: Ki=1.0 μM , mixed type, α=1.9; at pH 5.5: Ki=0.63 μM , mixed type, α=17). The N‐unsubstituted D ‐manno‐isoquinuclidine (+)‐ 3 is a poor inhibitor. Its inhibition strength and type also depend on the pH (at pH 4.5: Ki=1.2?103 μM , mixed type, α=1.1; at pH 5.5: Ki=0.25?103 μM , mixed type, α=11). The enantiomeric N‐benzyl L ‐manno‐isoquinuclidine (?)‐ 2 is a good inhibitor of snail β‐mannosidase, albeit noncompetitive (at pH 4.5: Ki=69 μM ). The N‐unsubstituted isoquinuclidine (?)‐ 2 is a poor inhibitor (at pH 4.5: IC50=7.3?103 μM ). A comparison of the inhibition by the pure manno‐isoquinuclidines (+)‐ 2 and (+)‐ 3 , (+)‐ 2 /(?)‐ 2 1 : 1, and (+)‐ 3 /(?)‐ 3 1 : 1 with the published data for racemic 2 and 3 led to a rectification of the published data. The inhibition of snail β‐mannosidase by the isoquinuclidines 2 and 3 suggests that the hydrolysis of β‐D ‐mannopyranosides by snail β‐mannosidase proceeds via a distorted conformer, in agreement with the principle of stereoelectronic control. 相似文献
A comprehensive method for the construction of a high‐mannose‐type glycan library by systematic chemo‐enzymatic trimming of a single Man9‐based precursor was developed. It consists of the chemical synthesis of a non‐natural tridecasaccharide precursor, the orthogonal demasking of the non‐reducing ends, and trimming by glycosidases, which enabled a comprehensive synthesis of high‐mannose‐type glycans in their mono‐ or non‐glucosylated forms. It employed glucose, isopropylidene, and N‐acetylglucosamine groups for blocking the A‐, B‐, and C‐arms, respectively. After systematic trimming of the precursor, thirty‐seven high‐mannose‐type glycans were obtained. The power of the methodology was demonstrated by the enzymatic activity of human recombinant N‐acetylglucosaminyltransferase‐I toward M7–M3 glycans, clarifying the substrate specificity in the context of high‐mannose‐type glycans. 相似文献
It was shown that retaining β‐glucosidases and galactosidases of families 1–3 feature a strong interaction between C(2)OH of the substrate and the catalytic nucleophile. An analogous interaction can hardly take place for retaining β‐mannosidases. A structure? activity comparison between the inhibition of the β‐glucosidase from Caldocellum saccharolyticum (family 1) and β‐glucosidase from sweet almonds by the gluco‐imidazoles 1 – 6 , and the inhibition of snail β‐mannosidase by the corresponding manno‐imidazoles 8 – 13 does not show any significant difference, suggesting that also the mechanisms of action of these glycosidases do not differ significantly. For this comparison, we synthesized and tested the manno‐imidazoles 9 – 13, 28, 29, 32, 35, 40, 41, 43, 46, 47 , and 50 . Among these, the alkene 29 is the strongest known inhibitor of snail β‐mannosidase (Ki=6 nM , non‐competitive); the aniline 35 is the strongest competitive inhibitor (Ki=8 nM ). 相似文献
The selective activation of the immune system using nanoparticles as a drug delivery system is a promising field in cancer therapy. Block copolymers from HPMA and laurylmethacrylate‐co‐hymecromone‐methacrylate allow the preparation of multifunctionalized core‐crosslinked micelles of variable size. To activate dendritic cells (DCs) as antigen presenting cells, the carbohydrates mannose and trimannose are introduced into the hydrophilic corona as DC targeting units. To activate DCs, a lipophilic adjuvant (L18‐MDP) is incorporated into the core of the micelles. To elicit an immune response, a model antigen peptide (SIINFEKL) is attached to the polymeric nanoparticle—in addition—via a click reaction with the terminal azide. Thereafter, the differently functionalized micelles are chemically and biologically characterized. While the core‐crosslinked micelles without carbohydrate units are hardly bound by DCs, mannose and trimannose functionalization lead to a strong binding. Flow cytometric analysis and blocking studies employing mannan suggest the requirement of the mannose receptor and DC‐SIGN for effective micelle binding. It could be suppressed by blocking with mannan. Adjuvant‐loaded micelles functionalized with mannose and trimannose activate DCs, and DCs preincubated with antigen‐conjugated micelles induce proliferation of antigen‐specific CD8+ T cells. 相似文献
A convenient and divergent approach was developed to prepare diverse bacterial 3‐deoxy‐d ‐manno‐oct‐2‐ulosonic acid (Kdo) oligosaccharides containing a Kdo‐α‐(2→4)‐Kdo fragment. The orthogonal protected α‐(2→4) linked Kdo‐Kdo disaccharide 3 , serving as a common precursor, was divergently transformed into the corresponding 8‐, 8′‐, and 4′‐hydroxy disaccharides 5 , 7 , and 14 , respectively. Then, these alcohols were glycosylated, respectively, with the 5,7‐O‐di‐tert‐butylsilylene (DTBS) protected Kdo thioglycoside donors 1 or 2 in an α‐stereoselective and high‐yielding manner to afford a range of Kdo oligosaccharides. Finally, removal of all protecting groups of the newly formed glycosides resulted in the desired free Kdo oligomer. 相似文献
The application of chemical‐modified gold nanoparticles (GNPs) as chiral selector for the enantioseparation based on pseudostationary phase‐CEC (PSP‐CEC) is presented. GNPs modified by thiolated β‐CD were characterized by NMR and FT‐IR. The nanoparticle size was determined to be of 9.5 nm (+2.5 nm) by Transmission Electron Microscopy (TEM) and UV spectra. Four pairs of dinitrophenyl‐labeled amino acid enantiomers (DL‐Val, Leu, Glu and Asp) and three pairs of drug enantiomers (RS‐chlorpheniramine, zopiclone and carvedilol) were analyzed by using modified GNPs as the chiral selector in PSP‐CEC. Good theoretical plate number (up to 2.4×105per meter) and separation resolution (up to 4.7) were obtained even with low concentration of modified GNPs (0.8–1.4 mg/mL). The corresponding concentration of β‐CD in the buffer was only 0.30?0.53 mM, which was much lower than the optimum concentration of 15 mM if pure β‐CD was used as chiral selector. Our results showed that thiolated β‐CD modified GNPs have more sufficient interaction with the analytes, resulting in significant enhancement of enantioseparation. The study shed light on potential usage of chemical modified GNPs as chiral selector for enantioseparation based on PSP‐CEC. 相似文献