Monika Poláková

A chemical synthesis of two novel phenyl glycosides of trisaccharides related to acetylarabinoxylan is described. The trisaccharides bear acetyl and arabinofuranosyl moieties at the non‐reducing‐end xylopyranosyl residue, which is substituted at positions 2 and 3. Both compounds were treated with various xylan deacetylases classified in different carbohydrate esterase (CE) families and significant differences between the families were found. While the arabinosylation hampers deacetylation by CE2‐CE5 and CE12 family members, both epitopes are deesterified by CE1 and in particular CE6 enzymes. The 3‐O‐acetylated 2‐O‐arabinosylated compound is also processed by CE7 and majority of CE16 esterases, but not by a hitherto non‐classified Flavobacterium johnsoniae acetylxylan esterase. The data suggests that a slow deesterification of the 2‐O‐acetylated 3‐O‐arabinosylated compound may be due to the acetyl group migration followed by deacetylation of this migration product.

Paenibacillus larvae is the causative agent of American foulbrood (AFB), the most serious bacterial disease affecting developing honeybee larvae and pupas. In this study, a library of 24 (thio)glycosides, glycosyl sulfones, 6-O-esters, and ethers derived from d-mannose, d-glucose, and d-galactose having C10 or C12 alkyl chain were evaluated for their antibacterial efficacy against two P. larvae strains. The efficacy of the tested compounds determined as minimal inhibitory concentrations (MICs) varied greatly. Generally, dodecyl derivatives were found to be more potent than their decylated analogs. Thioglycosides were more efficient than glycosides and sulfones. The activity of the 6-O-ether derivatives was higher than that of their ester counterparts. Seven derivatives with dodecyl chain linked (thio)glycosidically or etherically at C-6 showed high efficacy against both P. larvae strains (MICs ranged from 12.5 μM to 50 μM). Their efficacies were similar or much higher than those of selected reference compounds known to be active against P. larvae—lauric acid, monolaurin, and honeybee larval food components, 10-hydroxy-2-decenoic acid, and sebacic acid (MICs ranged from 25 μM to 6400 μM). The high efficacies of these seven derivatives suggest that they could increase the anti-P. larvae activity of larval food and improve the resistance of larvae to AFB disease through their application to honeybee colonies.

A synthesis of 1,4-imino-ᴅ-lyxitols and their N-arylalkyl derivatives altered at C-5 is reported. Their inhibitory activity and selectivity toward four GH38 α-mannosidases (two Golgi types: GMIIb from Drosophila melanogaster and AMAN-2 from Caenorhabditis elegans, and two lysosomal types: LManII from Drosophila melanogaster and JBMan from Canavalia ensiformis) were investigated. 6-Deoxy-DIM was found to be the most potent inhibitor of AMAN-2 (K_i = 0.19 μM), whose amino acid sequence and 3D structure of the active site are almost identical to the human α-mannosidase II (GMII). Although 6-deoxy-DIM was 3.5 times more potent toward AMAN-2 than DIM, their selectivity profiles were almost the same. N-Arylalkylation of 6-deoxy-DIM resulted only in a partial improvement as the selectivity was enhanced at the expense of potency. Structural and physicochemical properties of the corresponding inhibitor:enzyme complexes were analyzed by molecular modeling.

A binding mechanism of selective inhibitors of Golgi α-mannosidase II was elucidated by pK_aand FMO-PIEDA calculations. SyntheticN-substituted imino-d-lyxitol inhibitors were evaluated with four enzymes from the glycoside hydrolase GH38 family.

N-Alkylated 1,4-dideoxy-1,4-imino-L-lyxitols (N-alkylated polyhydroxypyrrolidines) differing in the alkyl chain length (from C7 to C14) and their capping functional group (methyl, amine, amidine and guanidine) have been synthesized. Based on molecular modeling, the structures were designed as selective inhibitors of Golgi α-mannosidase II (GMIIb) with a predicted reduced binding affinity toward lysosomal α-mannosidase II (LManII). In general, the potency of the derivatives toward GMIIb decreased in the following order: amidines > guanidines > alkanes > amines. The most potent inhibitors of GMIIb were N-nonylamidine and N-decylamidine 1,4-dideoxy-1,4-imino-L-lyxitols [K_i (GMIIb) = 4.0 μM and 5.5 μM]. Elongation of the alkyl chain led to an increase in potency while maintaining the selectivity toward the target enzyme. Molecular docking indicates that the alkyl chain length is crucial for an efficient interaction of the terminal group with a polar dyad (Asp270–Asp340) in the case of amidine, guanidine and amine structures and with a hydrophobic pocket (Pro298–Trp299) in the case of alkyl derivatives. Quantum mechanics calculations on conformational analysis of selected inhibitors indicate the differences in preferred envelope conformations for neutral and protonated forms of the pyrrolidine ring in a polar solution. NMR measurements in the polar solution at different pH values revealed that the N-substituted pyrrolidines with the terminal basic group may actually exhibit a different protonation form of the pyrrolidine ring under pH conditions occurring in Golgi apparatus compared with swainsonine or other N-substituted pyrrolidines lacking the terminal basic group.