Síntesis de fucooligosacáridos empleando α-L-fucosidasas microbianas: estudio in vitro e in silico
Some of the benefits provided by human milk to infants are attributed to the oligosaccharides it contains. Most of these oligosaccharides are fucosylated and mainly produce a prebiotic action, promoting the selective growth of bifidobacteria in the gastrointestinal tract, which is why there is inter...
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| Tipo de recurso: | tesis doctoral |
| Estado: | Versión publicada |
| Fecha de publicación: | 2023 |
| País: | México |
| Institución: | Universidad Autónoma Metropolitana |
| Repositorio: | Repositorio Institucional de la UAM Iztapalapa |
| Idioma: | español |
| OAI Identifier: | oai:bindani.izt.uam.mx:4q77fr86n |
| Acceso en línea: | https://doi.org/10.24275/uami.4q77fr86n |
| Access Level: | acceso abierto |
| Palabra clave: | info:eu-repo/classification/LEM/Enzymes -- Synthesis info:eu-repo/classification/LEM/Enzimas -- Síntesis info:eu-repo/classification/LEM/Oligosacáridos -- Síntesis info:eu-repo/classification/LEM/Oligosaccharides -- Sythesis info:eu-repo/classification/cti/6 |
| Sumario: | Some of the benefits provided by human milk to infants are attributed to the oligosaccharides it contains. Most of these oligosaccharides are fucosylated and mainly produce a prebiotic action, promoting the selective growth of bifidobacteria in the gastrointestinal tract, which is why there is interest in synthesizing them to take advantage of their biological activity. One of the methods available to synthesize them is enzymatic synthesis using glycosyl hydrolases such as α-L-fucosidase. The fucosidases belonging to the GH29 family are those that have shown greater ability to carry out the transfucosylation reaction. In the present work, an in silico study was carried out to study the differences between two α-L-fucosidases from the GH29A subfamily belonging to Thermotoga maritima and Lacticaseibacillus casei, as well as one of the GH29B subfamily belonging to Bifidobacterium longum subsp. infantis, identifying differences in the active site that are related to the affinity with the fucosyl donor substrate, para-nitrophenyl fucose. Furthermore, a simulation of the synthesis reaction of fucosylated oligosaccharides was carried out by molecular docking with the α-L-fucosidase from T. maritima, finding favorable binding sites for the acceptor substrate lactose, that are occupied before the substrate approach the active site. Subsequently, the interaction between the α-L-fucosidase from B. longum subsp. infantis with the donor substrates 2'- fucosyl lactose and 3-fucosyl lactose were examinated, as well with lactose and lactulose as acceptor substrates, verifying that it is theoretically possible to carry out the transfucosylation reaction with this enzyme. Finally, the synthesis of fucosylated oligosaccharides was carried out in vitro, employing an B. longum enzymatic extract, and using para-nitrophenyl fucose as a fucosyl donor substrate and lactose, lactulose, galactose and fructose as acceptor substrates, obtaining yields of 0.046%, 0.16%. 0.32% and 2.4% respectively, confirming the transfer of fucose to fructose through the acid hydrolysis of the fucose-fructose disaccharide, obtaining an equimolar proportion of 0.9: 1. Keywords: Bifidobacteria, α-L-fucosidase, fucooligosaccharides, transfucosylation. |
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