A comparative study of the substrate preference of the sialidases, CpNanI, HpNanH, and BbSia2 towards 2-Aminobenzamide-labeled 3'-Sialyllactose, 6'-Sialyllactose, and Sialyllacto-N-tetraose-b.

Sialidases catalyze the removal of terminal sialic acids from sialylated biomolecules, and their substrate preference is frequently indicated in terms of the glycosidic linkages cleaved (α2-3, α2-6, and α2-8) without mention of the remaining sub-terminal reducing-end saccharide moieties. Many human gut commensal and pathogenic bacteria secrete sialidases to forage for sialic acids, which are then utilized as an energy source or assimilated into membrane/capsular structural components. Infant gut commensals similarly utilize sialylated human milk oligosaccharides containing different glycosidic linkages. Here, we have studied the preference of the bacterial sialidases, BbSia2 from Bifidobacterium bifidum, CpNanI from Clostridium perfringens, and HpNanH from Glaesserella parasuis, for the glycosidic linkages, Siaα2-3Gal, Siaα2-6Gal, and Siaα2-6GlcNAc, by employing 2-Aminobenzamide-labeled human milk oligosaccharides, 3'-Sialyllactose (3'-SL), 6'-Sialyllactose (6'-SL), and Sialyllacto-N-tetraose-b (LSTb), respectively, as proxies for these glycosidic linkages. BbSia2, CpNanI, and HpNanH hydrolyzed these three oligosaccharides with the glycosidic linkage preferences, 3'-SL (Siaα2-3Gal) ≥ LSTb (Siaα2-6GlcNAc) ≥ 6'-SL (Siaα2-6Gal), 3'-SL (Siaα2-3Gal) ≥ 6'-SL (Siaα2-6Gal) > LSTb (Siaα2-6GlcNAc), and 3'-SL (Siaα2-3Gal) ≥ 6'-SL (Siaα2-6Gal) > LSTb (Siaα2-6GlcNAc), respectively. Our finding suggests that sub-terminal reducing-end saccharide moieties can profoundly influence the substrate preference of sialidases, and advocates for the characterization and indication of the substrate preference of sialidases in terms of both the glycosidic linkage and the sub-terminal reducing-end saccharide moiety.

Surface activity, kinetics, thermodynamics and comparative study of adsorption of selected cationic and anionic dyes onto H3PO4-functionalized bagasse from aqueous stream.

The discharge of dyes into the water body creates toxicity to aquatic organisms because of their aromatic structure and difficult degradation. So, the treatment of dye-contaminated wastewater is required before releasing it. In the present study, thermally treated (600 °C) and H3PO4 (55%)-functionalized bagasse, henceforth called thermochemically activated bagasse (TCAB), was synthesized as potential adsorbent for the effective removal of selected cationic and anionic dyes from their aqueous stream. TCAB characterization was done employing FT-IR, SEM, XRD, zeta potential, BET, and PZC techniques. The comparative study shows that the relative adsorption on TCAB followed the sequence, methyl red (185 mg/g) > safranin (178 mg/g) > congo red (146 mg/g) > brilliant green (139 mg/g) > malachite green (130 mg/g) > bromocresol green (94 mg/g). The adsorption efficiency was investigated concerning the effect of change in TCAB dose (0.05-0.3 g/100 mL), initial dye concentration (20-200 mg/L), pH (4.0-10.0), ionic strength (0.1-0.5 M KCl), urea concentration (0.1-0.5 M) and temperature (25-45 °C). The representative adsorption isotherms belong to typical L-type. The time-dependent dye removal was best explained by the pseudo-second-order (PSO) kinetic model (R2 = 0.9859-0.9991), while equilibrium data were best explained by the Freundlich model (R2 = 0.9881-0.9961). Thermodynamic study showed the spontaneous (ΔG0 <0) and exothermic nature (ΔH0 <0) of the adsorption of different cationic and anionic dyes. The cyclic adsorption ability of TCAB for different dyes was checked up to three cycles (185 to 168 mg/L for MR, 178 to 165 mg/L for SF, 146 to 130 mg/L for CR, 139 to 127 mg/L for BG, 130 to 114 mg/L for MG and 94 to 80 mg/L for BCG), and no significant decrease in the adsorption capacity was noticed. So, the present study provides valuable insights into the adsorption of cationic and anionic dyes onto H3PO4-functionalized bagasse. Addressing the adsorptive aspects enhances the clarity, reliability and applicability of the study's findings and contributes to its overall scientific impact.

New carbohydrate binding domains identified by phage display based functional metagenomic screens of human gut microbiota.

Uncultured microbes represent a huge untapped biological resource of novel genes and gene products. Although recent genomic and metagenomic sequencing efforts have led to the identification of numerous genes that are homologous to existing annotated genes, there remains, yet, an enormous pool of unannotated genes that do not find significant sequence homology to existing annotated genes. Functional metagenomics offers a way to identify and annotate novel gene products. Here, we use functional metagenomics to mine novel carbohydrate binding domains that might aid human gut commensals in adherence, gut colonization, and metabolism of complex carbohydrates. We report the construction and functional screening of a metagenomic phage display library from healthy human fecal samples against dietary, microbial and host polysaccharides/glycoconjugates. We identify several protein sequences that do not find a hit to any known protein domain but are predicted to contain carbohydrate binding module-like folds. We heterologously express, purify and biochemically characterize some of these protein domains and demonstrate their carbohydrate-binding function. Our study reveals several previously unannotated carbohydrate-binding domains, including a levan binding domain and four complex N-glycan binding domains that might be useful for the labeling, visualization, and isolation of these glycans.