Synthesis and Evaluation as Glycosidase Inhibitors of Isoquinuclidines Mimicking a Distorted β‐Mannopyranoside

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: K_i=1.0 μM , mixed type, α=1.9; at pH 5.5: K_i=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: K_i=1.2?10³ μM , mixed type, α=1.1; at pH 5.5: K_i=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: K_i=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.     

Norbornane Mimics of Distorted β‐D‐Glucopyranosides – Inhibitors of β‐D‐Glucopyranosidases?

The racemic gluco‐configured norbornanes 4 and 16 were prepared and tested as inhibitors of β‐glucosidases. The known alcohol 5 was deprotected to provide the triol 6 . Silylation (→ 7 ), monobenzoylation (→ 8 / 9 ), and oxidation provided the regioisomeric ketones 10 and 11 . Reduction of 10 gave the desired endo‐alcohol 13 , albeit in low yield, while reduction of the isomeric ketone 11 provided mostly the altro‐configured endo‐alcohol 12 . The alcohol 13 was desilylated to 14 . Debenzoylation to 15 followed by hydrogenolytic deprotection gave the amino triol 4 that was reductively aminated to the benzylamine 16 . The amino triols 4 and 16 proved weak inhibitors of the β‐glucosidase from Caldocellum saccharolyticum ( 4 : IC₅₀ = 5.6 mm; 16 : IC₅₀ = 3.3 mm) and from sweet almonds ( 16 : IC₅₀ = 5.5 mm) . A comparison of 4 with the manno‐configured norbornane 3 shows that 3 is a better inhibitor of snail β‐mannosidase than 4 is of β‐glucosidases, in keeping with earlier results suggesting that these β‐glycosidases enforce a different conformational itinerary.     

Synthesis of Tetrahydropyridoimidazole‐2‐acetates: Effect of Carboxy and Methoxycarbonyl Groups at C(2) on the Inhibition of Some β‐ and α‐Glycosidases

The gluco‐ and manno‐tetrahydropyridoimidazole‐2‐acetates and ‐acetic acids 16 and 17 , and 20 and 21 , respectively, were synthesized by condensation, in the presence of HgCl₂, of the known thionolactam 26 with the β‐amino ester 25 that was obtained by addition of AcOMe to the imine 22 , followed by debenzylation. The resulting methyl esters 16 and 20 were hydrolyzed to the acetic acids 17 and 21 . The (methoxycarbonyl)‐imidazole 14 and the acid 15 were obtained via the known aldehyde 29 . The imidazoles 14 – 17, 20 , and 21 were tested as inhibitors of the β‐glucosidase from Caldocellum saccharolyticum, the α‐glucosidase from brewer's yeast, the β‐mannosidase from snail, and the α‐mannosidase from Jack beans (Tables 1–3). There is a similar dependence of the K_i values on the nature of the C(2)‐substituent in the gluco‐ and manno‐series. With the exception of 19 , manno‐imidazoles are weaker inhibitors than the gluco‐analogues. The methyl acetates 16 and 20 are 3–4 times weaker than the methyl propionates 5 and 11 , in agreement with the hydrophobic effect. The gluco‐configured (methoxycarbonyl)‐imidazole 14 is 20 times weaker than the methyl acetate 16 , reflecting the reduced basicity of 14 , while the manno‐configured (methoxycarbonyl)‐imidazole 18 is only 1.2 times weaker than the methyl acetate 20 , suggesting a binding interaction of the MeOCO group and the β‐mannosidase. The carboxylic acids 6, 12, 15, 17, 19 , and 21 are weaker inhibitors than the esters, with the propionic acids 6 and 12 being the strongest and the carboxy‐imidazoles 15 and 19 the weakest inhibitors. The manno‐acetate 21 inhibits the β‐mannosidase ca. 8 times less strongly than the propionate 12 , but only 1.5 times more strongly than the carboxylate 19 , suggesting a compensating binding interaction also of the COOH group and the β‐mannosidase. The α/β selectivity for the gluco‐imidazoles ranges between 110 for 15 and 13.4?10³ for 6 ; the manno‐imidazoles are less selective. The methyl propionates proved the strongest inhibitors of the α‐glucosidase (IC₅₀ ( 5 )=25 μM ) and the α‐mannosidase (K_i( 11 ) =0.60 μM ).     

Synthesis of C(2)‐Substituted manno‐Configured Tetrahydroimidazopyridines and Their Evaluation as Inhibitors of Snail β‐Mannosidase

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 (K_i=6 nM , non‐competitive); the aniline 35 is the strongest competitive inhibitor (K_i=8 nM ).     

Probing the Interaction of the Hydroxy Group at C(4) of Lactone‐Type Inhibitors with β‐Glucosidases and β‐Galactosidases

The inhibition of the β‐glucosidases from sweet almonds and from Caldocellum saccharolyticum by the 4‐amino‐4‐deoxy lactam 11 , the 4‐deoxy lactam 12 , and the corresponding imidazoles 13 and 14 was compared to the inhibition by the hydroxy analogues 1 and 3 . Substitution of the OH group at C(4) by an amino group or by hydrogen weakened the inhibition by ΔΔG_diss = + 1.9 to + 3.1 kcal/mol. Similarly, the inhibition of the β‐galactosidase from bovine liver and from E. coli by the 4‐deoxy lactam 12 and the imidazole 14 , as compared to the one by the galacto‐configured lactam 9 and imidazole 10 , is weakened by deoxygenation at C(4) (ΔΔG_diss = + 2.6 and 4.5 kcal/mol, resp.). The effect of these substitutions on the inhibition of the C. saccharolyticum β‐glucosidase is slightly stronger than the one on the sweet almonds β‐glucosidases. The effect is also stronger on the inhibition by the imidazoles than by the lactams, and depends on the flexibility of the inhibitors. The amino and deoxy lactams 11 and 12 were prepared from the galactonolactam‐derived triflate 17 by substitution with azide and hydride, respectively, followed by hydrogenation. Azidation of the galacto‐configured imidazopyridine‐derived triflate 24 and hydrogenation gave the amino‐imidazole 13 . The deoxy lactam 20 was transformed to the manno‐ and gluco‐configured deoxy‐imidazoles 29 and 30 via the thionolactam 28 . Hydrogenolytic deprotection of 30 gave the deoxy‐imidazole 14 .     

Synthesis of New C(2)‐Substituted gluco‐Configured Tetrahydroimidazopyridines and Their Evaluation as Glucosidase Inhibitors

The gluco‐configured C(2)‐substituted tetrahydroimidazopyridines 8 – 14 were prepared and tested as inhibitors of the β‐glucosidases from Caldocellum saccharolyticum and from sweet almonds, and of the α‐glucosidase from brewer's yeast. All new imidazopyridines are nanomolar inhibitors of the β‐glucosidases and micromolar inhibitors of the α‐glucosidase. The 3‐phenylpropyl derivative 14 proved the strongest inhibitor of the Caldocellum β‐glucosidase (K_i = 0.9 nM ), only slightly weaker than the known 2‐phenylethyl analogue 7 , and the propyl derivative 13 is the strongest inhibitor of the sweet almond β‐glucosidases (K_i = 3.2 nM ), again slightly weaker than 7 . There is no strong dependence of the inhibition on the nature of the C(2)‐substituent and no clear correlation between the inhibitory strength of the known manno‐configured imidazopyridines 2 – 6 and the gluco‐analogues 8 – 12 . While most manno‐imidazopyridines are competitive inhibitors, the gluco‐analogues proved non‐competitive inhibitors of the Caldocellum β‐glucosidase and mixed‐type or partial mixed‐type inhibitors of the sweet almond β‐glucosidases.     

Improved Access to Imidazole‐phosphonic Acids: Synthesis of D‐manno‐Tetrahydroimidazopyridine‐2‐phosphonates

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).     

Synthesis of Glycaro‐1,5‐lactams and Tetrahydrotetrazolopyridine‐5‐carboxylates: Inhibitors of β‐D‐Glucuronidase and α‐L‐Iduronidase

The known glucaro‐1,5‐lactam 8 , its diastereoisomers 9 – 11 , and the tetrahydrotetrazolopyridine‐5‐carboxylates 12 – 14 were synthesised as potential inhibitors of β‐D ‐glucuronidases and α‐L ‐iduronidases. The known 2,3‐di‐O‐benzyl‐4,6‐O‐benzylidene‐D ‐galactose ( 16 ) was transformed into the D ‐galactaro‐ and L ‐altraro‐1,5‐lactams 9 and 11 via the galactono‐1,5‐lactam 21 in twelve steps and in an overall yield of 13 and 2%, respectively. A divergent strategy, starting from the known tartaric anhydride 41 , led to the D ‐glucaro‐1,5‐lactam 8 , D ‐galactaro‐1,5‐lactam 9 , L ‐idaro‐1,5‐lactam 10 , and L ‐altraro‐1,5‐lactam 11 in ten steps and in an overall yield of 4–20%. The anhydride 41 was transformed into the L ‐threuronate 46 . Olefination of 46 to the (E)‐ or (Z)‐alkene 47 or 48 followed by reagent‐ or substrate‐controlled dihydroxylation, lactonisation, azidation, reduction, and deprotection led to the lactams 8 – 11 . The tetrazoles 12 – 14 were prepared in an overall yield of 61–81% from the lactams 54, 28 , and 67 , respectively, by treatment with Tf₂O and NaN₃, followed by saponification, esterification, and hydrogenolysis. The lactams 8 – 11 and 40 and the tetrazoles 12 – 14 are medium‐to‐strong inhibitors of β‐D ‐glucuronidase from bovine liver. Only the L ‐ido‐configured lactam 10 (K_i = 94 μM ) and the tetrazole 14 (K_i = 1.3 mM ) inhibit human α‐L ‐iduronidase.     

Synthesis of Glucuronic,Mannuronic, and Galacturonic Acid‐Derived Imidazoles as Inhibitors of Bovine Liver β‐Glucuronidase

The gluco‐, manno‐, and galacto‐configured imidazopyridine‐5‐carboxylates 5 – 7 , respectively, were synthesized and evaluated as inhibitors of bovine liver β‐glucuronidase. The gluconolactam 15 was transformed into the gluco‐ and manno‐imidazoles 5 and 6 in nine steps and in an overall yield of 9 and 12%, respectively. Oxidation and esterification of the selectively protected gluco‐ and manno‐configured hydroxymethyl‐imidazopyridines 23 and 25 , respectively (both obtained from gluconolactam 15 ), provided the benzhydryl esters 24 and 26 , respectively. Hydrogenolysis afforded the gluco‐imidazopyridine‐carboxylic acid 5 and the manno‐isomer 6 . Similarly, the hydroxymethyl‐imidazopyridine 33 , obtained from galactonolactam 27 , was subjected to oxidation, esterification, and deprotection to afford the galacto‐configured imidazopyridine‐carboxylate 7 in ten steps from the galactonolactam 27 and in an overall yield of 13%. The gluco‐configured imidazole 5 is the strongest known inhibitor of β‐glucuronidases (K_i = 12 nM ), while the manno‐ and galacto‐configured imidazoles 6 and 7 are micromolar inhibitors of bovine β‐glucuronidase. The small difference between the inhibitory strength of the imidazopyridine‐carboxylic acid 5 and the tetrazolopyridine‐carboxylic acid 1 , and the difference between the configurational selectivity of 5 – 7 as compared to the unselectivity of the corresponding lactams 3 and 4 are discussed.     

Chemo‐Enzymatic Synthesis of 3‐Deoxy‐β‐D‐ribofuranosyl Purines

9‐(3‐Deoxy‐β‐D ‐erythro‐pentofuranosyl)‐2,6‐diaminopurine ( 6 ) was synthesized by an enzymatic transglycosylation of 2,6‐diaminopurine ( 2 ) with 3′‐deoxycytidine ( 1 ) as a donor of 3‐deoxy‐D ‐erythro‐pentofuranose moiety. This transformation comprises i) deamination of 1 to 3′‐deoxyuridine ( 3 ) under the action of whole cell (E. coli BM‐11) cytidine deaminase (CDase), ii) the phosphorolytic cleavage of 3 by uridine phosphorylase (UPase) giving rise to the formation of uracil ( 4 ) and 3‐deoxy‐α‐D ‐erythro‐pentofuranose‐1‐O‐phosphate ( 5 ), and iii) coupling of the latter with 2 catalyzed by whole cell (E. coli BMT‐4D/1A) purine nucleoside phosphorylase (PNPase). Deamination of 6 by adenosine deaminase (ADase) gave 3′‐deoxyguanosine ( 7 ). Treatment of 6 with NaNO₂ afforded 9‐(3‐deoxy‐β‐D ‐erythro‐pentofuranosyl)‐2‐amino‐6‐oxopurine (3′‐deoxyisoguanosine; 8 ). Schiemann reaction of 6 (HF/HBF₄+NaNO₂) gave 9‐(3‐deoxy‐β‐D ‐erythro‐pentofuranosyl)‐2‐fluoroadenine ( 9 ).     

Syntheses of β‐C(1→3)‐Glucopyranosides of 2‐ and 4‐Deoxy‐D‐hexoses

The Oshima? Nozaki (Et₂AlI) condensation of isolevoglucosenone ( 4 ) with 2,6‐anhydro‐3,4,5,7‐tetra‐O‐benzyl‐D ‐glycero‐D ‐gulo‐heptose ( 5 ) gave an enone 6 that was converted with high stereoselectivity to 3‐C‐[(1R)‐2,6‐anhydro‐D ‐glycero‐D ‐gulo‐heptitol‐1‐C‐yl]‐2,3‐dideoxy‐D ‐arabino‐hexose ( 1 ; 1 : 1 mixture of α‐ and β‐D ‐pyranose), and to 3‐C‐[(1R)‐2,6‐anhydro‐D ‐glycero‐D ‐gulo‐heptitol‐1‐C‐yl]‐2,3‐dideoxy‐D ‐lyxo‐hexose ( 2 ; 2.7 : 1.4 : 1.0 : 1.4 mixture of α‐D ‐furanose, β‐D ‐furanose, α‐D ‐pyranose, and β‐D ‐pyranose). The Oshima? Nozaki (Et₂AlI) condensation of levoglucosenone ( 17 ) with aldehyde 5 gave an enone 18 that was converted with high stereoselectivity to 3‐C‐[(1R)‐2,6‐anhydro‐D ‐glycero‐D ‐gulo‐heptitol‐1‐C‐yl]‐3,4‐dideoxy‐α‐D ‐arabino‐hexopyranose ( 3 ; single anomer).     

Synthesis of 5′‐O‐(2‐Azido‐2‐deoxy‐α‐D‐glycosyl)nucleosides and Their Antitumor Activities

The 1,3,4,6‐tetra‐O‐acetyl‐2‐azido‐2‐deoxy‐β‐D ‐mannopyranose ( 4 ) or the mixture of 1,3,6‐tri‐O‐acetyl‐2‐azido‐2‐deoxy‐4‐O‐(2,3,4,6‐tetra‐O‐acetyl‐β‐D ‐galactopyranosyl)‐β‐D ‐mannopyranose ( 10 ) and the corresponding α‐D ‐glucopyranose‐type glycosyl donor 9 / 10 reacted at room temperature with protected nucleosides 12 – 15 in CH₂Cl₂ solution in the presence of BF₃?OEt₂ as promoter to give 5′‐O‐(2‐azido‐2‐deoxy‐α‐D ‐glycosyl)nucleosides in reasonable yields (Schemes 2 and 3). Only the 5′‐O‐(α‐D ‐mannopyranosyl)nucleosides were obtained. Compounds 21, 28, 30 , and 31 showed growth inhibition of HeLa cells and hepatoma Bel‐7402 cells at a concentration of 10 μM in vitro.     

Sugar‐Derived Di‐ and Tetrahydropyridazinones: Synthesis of New Glycosidase Inhibitors

The N‐unsubstituted D ‐arabino‐tetrahydropyridazinone 7 is a micromolar inhibitor of β‐glucosidases from sweet almonds (competitive), Caldocellum saccharolyticum (mixed), yeast α‐glucosidase (competitive), jack bean α‐mannosidase (competitive), and snail β‐mannosidase (competitive). The N‐substituted tetrahydropyridazinones 22 , 24 , and 26 are weak inhibitors of these glycosidases, and so are the dihydropyridazinones 8 and 17 – 19 , where the best inhibition was observed for 8 (K_i=56 μM for jack bean α‐mannosidase). The tetrahydropyridazinones were obtained by reduction of the corresponding dihydropyridazinones with NaCNBH₃, and the dihydropyridazinones were prepared by treatment with hydrazine or substituted hydrazines of the known and readily available D ‐threo‐pent‐2‐uluronate 11 .     

Probing the Conformational Changes in the Enzymatic Hydrolysis of 2‐Acetamido‐2‐deoxy‐β‐D‐glucopyranosides

The isoquinuclidines 7 and 8 were synthesised and tested as inhibitors of hexosaminidases from jack beans and from bovine kidney. These isoquinuclidines mimick the ^1,4B‐conformer of a N‐acetyl‐glucosamine‐derived β‐d‐ glucopyranoside; they are competitive inhibitors with K_i values from 0.014 to 0.30 μM . The strong inhibition of these enzymes agrees with the hypothesis that the enzymatic hydrolysis of 2‐acetamido‐2‐deoxy‐β‐d‐ glucopyranosides proceeds via a boat‐like conformer with a pseudo‐axial scissile glycosidic bond and a pseudo‐axial acetamido substituent optimally oriented to effect an intramolecular substitution of the aglycon.     

Cationic ring‐opening polymerization of 1,6‐anhydro‐2,3,4‐tri‐O‐allyl‐β‐D‐glucopyranose as a convenient synthesis of dextran

For the convenient synthesis of (1→6)‐α‐D ‐glucopyranan, i. e., dextran ( 4 ), ring‐opening polymerization of 1,6‐anhydro‐2,3,4‐tri‐O‐allyl‐β‐D ‐glucopyranose ( 1 ) has been carried out using BF₃·OEt₂. With a ratio of [BF₃·OEt₂]/[ 1 ] = 0.5 at 0 °C for 140 h, the yield and M_n of the obtained polymer are 84.0% and 21 700, respectively. The polymer consists of (1→6)‐α‐linked 2,3,4‐tri‐O‐allyl‐D ‐glucopyranose ( 2 ) which is similar to the results for the cationic ring‐opening polymerization of 1,6‐anhydro‐2,3,4‐tri‐O‐methyl‐β‐D ‐glucopyranose and 1,6‐anhydro‐2,3,4‐tri‐O‐ethyl‐β‐D ‐glucopyranose. Polymer 2 was isomerized using tris(triphenylphosphine)‐chlororhodium as the catalyst in toluene/ethanol/water to yield polymeric 2,3,4‐tri‐O‐propenyl‐(1→6)‐α‐D ‐glucopyranan ( 3 ). Deprotection of the propenyl ether linkage of 3 was then performed using hydrochloric acid in acetone to give 4 .     

Synthesis and Conformational Analysis of 1′‐ and 3′‐Substituted 2‐Deoxy‐2‐fluoro‐D‐ribofuranosyl Nucleosides

Convergent syntheses of the 9‐(3‐X‐2,3‐dideoxy‐2‐fluoro‐β‐D ‐ribofuranosyl)adenines 5 (X=N₃) and 7 (X=NH₂), as well as of their respective α‐anomers 6 and 8 , are described, using methyl 2‐azido‐5‐O‐benzoyl‐2,3‐dideoxy‐2‐fluoro‐β‐D ‐ribofuranoside ( 4 ) as glycosylating agent. Methyl 5‐O‐benzoyl‐2,3‐dideoxy‐2,3‐difluoro‐β‐D ‐ribofuranoside ( 12 ) was prepared starting from two precursors, and coupled with silylated N⁶‐benzoyladenine to afford, after deprotection, 2′,3′‐dideoxy‐2′,3′‐difluoroadenosine ( 13 ). Condensation of 1‐O‐acetyl‐3,5‐di‐O‐benzoyl‐2‐deoxy‐2‐fluoro‐β‐D ‐ribofuranose ( 14 ) with silylated N²‐palmitoylguanine gave, after chromatographic separation and deacylation, the N⁷‐β‐anomer 17 as the main product, along with 2′‐deoxy‐2′‐fluoroguanosine ( 15 ) and its N⁹‐α‐anomer 16 in a ratio of ca. 42 : 24 : 10. An in‐depth conformational analysis of a number of 2,3‐dideoxy‐2‐fluoro‐3‐X‐D ‐ribofuranosides (X=F, N₃, NH₂, H) as well as of purine and pyrimidine 2‐deoxy‐2‐fluoro‐D ‐ribofuranosyl nucleosides was performed using the PSEUROT (version 6.3) software in combination with NMR studies.     

Solution Structure of a RNA Decamer Duplex,Containing 9‐[2‐O‐(β‐D‐ribofuranosyl)‐β‐D‐ribofuranosyl]adenine,a Special Residue in Lower Eukaryotic Initiator tRNAs

The solution structure of the self‐complementary deca‐ribonucleotide 5′‐r(GCGA*AUUCGC)‐3′ containing 9‐[2‐O‐(β‐D ‐ribofuranosyl)‐β‐D ‐ribofuranosyl]adenine (A*), a modified nucleotide that occurs in lower eukaryotic methionine initiator tRNAs (tRNAs_i^Met), was determined by NMR spectroscopy. Unexpectedly, the modification has no effect on the thermal stability of the duplex. However, the extra ribose moiety is in the C(3′)‐endo conformation and takes up a well‐defined position in the minor groove, which is in agreement with its position in tRNAs_i^Met as determined by X‐ray crystallography. Molecular‐dynamics simulations on the RNA duplex in H₂O show that the position of the extra ribofuranose moiety seems to be stabilized by bridged H‐bonds (mediated by two H₂O molecules) to the backbone of the complementary chain.     

Synthesis of β‐Peptides with β‐Helices from New C‐Linked Carbo‐β‐Amino Acids: Study on the Impact of Carbohydrate Side Chains

The design and synthesis of β‐peptides from new C‐linked carbo‐β‐amino acids (β‐Caa) presented here, provides an opportunity to understand the impact of carbohydrate side chains on the formation and stability of helical structures. The β‐amino acids, Boc‐(S)‐β‐Caa_(g)‐OMe 1 and Boc‐(R)‐β‐Caa_(g)‐OMe 2 , having a D ‐galactopyranoside side chain were prepared from D ‐galactose. Similarly, the homo C‐linked carbo‐β‐amino acids (β‐hCaa); Boc‐(S)‐β‐hCaa_(x)‐OMe 3 and Boc‐(R)‐β‐hCaa_(x)‐OMe 4 , were prepared from D ‐glucose. The peptides derived from the above monomers were investigated by NMR, CD, and MD studies. The β‐peptides, especially the shorter ones obtained from the epimeric (at the amine stereocenter C_β) 1 and 2 by the concept of alternating chirality, showed a much smaller propensity to form 10/12‐helices. This substantial destabilization of the helix could be attributed to the bulkier D ‐galactopyranoside side chain. Our efforts to prepare peptides with alternating 3 and 4 were unsuccessful. However, the β‐peptides derived from alternating geometrically heterochiral (at C_β) 4 and Boc‐(R)‐β‐Caa_(x)‐OMe 5 (D ‐xylose side chain) display robust right‐handed 10/12‐helices, while the mixed peptides with alternating 4 and Boc‐β‐hGly‐OMe 6 (β‐homoglycine), resulted in left‐handed β‐helices. These observations show a distinct influence of the side chains on helix formation as well as their stability.     

Selective Formation of β‐D‐Glucosides of Hindered Alcohols

Alternative conditions for the classical glycosidation method of Koenigs‐Knorr allowed us to prepare selectively β‐D ‐glucosides of several hindered alcohols in good yields in a weakly acidic to almost neutral medium. To illustrate the versatility of our conditions, we prepared the β‐D ‐glucoside of an acid‐sensitive aglycone, a key‐intermediate for the total synthesis of a natural cyanoglucoside, bauhinin.     

Oligosaccharide Analogues of Polysaccharides Part 25

The ethynylated gluco‐azide 11 was prepared from the dianhydrogalactose 7 by ethynylation, transformation into the dianhydromannose 10 , and opening of the oxirane ring by azide (Scheme 1). The retentive alkynylating ring opening of 11 and of the corresponding amine 12 failed. (2‐Acetamidoglucopyranosyl)acetylenes were, therefore, prepared from the corresponding mannopyranosylacetylenes. Retentive alkynylating ring opening of the partially protected β‐D ‐mannopyranose 15 , possessing a C(3) OH group, gave a 85 : 15 mixture of 16 and the (E)‐enyne 17 . The alkyne 16 was deprotected to the tetrol 18 that was selectively protected and transformed into the C(2) O triflate 20 . Treatment with NaN₃ in DMF afforded a 85 : 15 mixture of the β‐D ‐gluco configured azide 21 and the elimination product 22 . Similarly, the α‐D ‐mannopyranosylacetylene 23 was transformed into the azide 26 . Retentive alkynylating ring opening of the ethynylated anhydromannose 28 gave the expected β‐D ‐mannopyranosyl 1,4‐dialkyne 29 as the main product besides the diol 28 , the triol 31 , and the (E)‐enyne 30 (Scheme 2). This enyne was also obtained from 31 by a stereoselective carboalumination promoted by the cis (axial) HO C(2) group. Deprotection of the dialkynylated mannoside 31 led to 32 , whereas selective silylation, triflation, and azidation gave a 3 : 7 mixture of the 1‐ethynylglucal 35 and the β‐D ‐gluco azide 36 , which was transformed into the diethynylated β‐D ‐GlcNAc analogue 38 . Similarly, the diethynylated α‐D ‐mannopyranoside 39 was transformed into the disilylated α‐D ‐GlcNAc analogue 41 , and further into the diol 42 and the monosilyl ether 43 (Scheme 5). Eglinton coupling of 41 gave the symmetric buta‐1,3‐diyne 44 , which did not undergo any further Eglinton coupling, even under forcing conditions. However, Eglinton coupling of the monosilyl ether 43 and subsequent desilylation gave the C₁‐symmetric cyclotrimer 45 in moderate yields.